Hybrid maize 33M72

ABSTRACT

According to the invention, there is provided a hybrid maize plant, designated as 33M72, produced by crossing an inbred maize line with a Pioneer Hi-Bred International, Inc. proprietary inbred maize line. This invention relates to the hybrid seed 33M72, the hybrid plant produced from the seed, and variants, mutants, and trivial modifications of hybrid 33M72. This invention also relates to methods for producing a maize plant containing in its genetic material one or more transgenes and to the transgenic maize plants produced by those methods. This invention further relates to methods for producing maize lines derived from hybrid maize line 33M72 and to the maize lines derived by the use of those methods.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of and priority to provisional U.S.Patent Application No. 60/540,649 filed Jan. 30, 2004, the contents ofwhich are hereby incorporated by reference in their entirety.

FIELD OF THE INVENTION

This invention relates generally to the field of maize breeding,specifically relating to hybrid maize designated 33M72.

BACKGROUND OF THE INVENTION Plant Breeding

The goal of plant breeding is to combine in a single variety or hybridvarious desirable traits. For field crops, these traits may includeresistance to diseases and insects, tolerance to heat and drought,reducing the time to crop maturity, greater yield, and better agronomicquality. With mechanical harvesting of many crops, uniformity of plantcharacteristics such as germination and stand establishment, growthrate, maturity, and plant and ear height is important.

Field crops are bred through techniques that take advantage of theplant's method of pollination. A plant is self-pollinated if pollen fromone flower is transferred to the same or another flower of the sameplant. A plant is sib pollinated when individuals within the same familyor line are used for pollination. A plant is cross-pollinated if thepollen comes from a flower on a different plant from a different familyor line. The term “cross pollination” and “out-cross” as used herein donot include self pollination or sib pollination.

Plants that have been self-pollinated and selected for type for manygenerations become homozygous at almost all gene loci and produce auniform population of true breeding progeny. A cross between twodifferent homozygous lines produces a uniform population of hybridplants that may be heterozygous for many gene loci. A cross of twoplants each heterozygous at a number of gene loci will produce apopulation of heterogeneous plants that differ genetically and will notbe uniform.

Maize (Zea mays L.), often referred to as corn in the United States, canbe bred by both self-pollination and cross-pollination techniques. Maizehas separate male and female flowers on the same plant, located on thetassel and the ear, respectively. Natural pollination occurs in maizewhen wind blows pollen from the tassels to the silks that protrude fromthe tops of the ears.

The development of a hybrid maize variety in a maize plant breedingprogram involves three steps: (1) the selection of plants from variousgermplasm pools for initial breeding crosses; (2) the selfing of theselected plants from the breeding crosses for several generations toproduce a series of inbred lines, which, individually breed true and arehighly uniform; and (3) crossing a selected inbred line with anunrelated inbred line to produce the hybrid progeny (F1). After asufficient amount of inbreeding successive filial generations willmerely serve to increase seed of the developed inbred. Preferably, aninbred line should comprise homozygous alleles at about 95% or more ofits loci.

During the inbreeding process in maize, the vigor of the linesdecreases. Vigor is restored when two different inbred lines are crossedto produce the hybrid progeny (F1). An important consequence of thehomozygosity and homogeneity of the inbred lines is that the hybridcreated by crossing a defined pair of inbreds will always be the same.Once the inbreds that create a superior hybrid have been identified, acontinual supply of the hybrid seed can be produced using these inbredparents and the hybrid corn plants can then be generated from thishybrid seed supply.

Large scale commercial maize hybrid production, as it is practicedtoday, requires the use of some form of male sterility system whichcontrols or inactivates male fertility. A reliable method of controllingmale fertility in plants also offers the opportunity for improved plantbreeding. This is especially true for development of maize hybrids,which relies upon some sort of male sterility system. There are severalways in which a maize plant can be manipulated so that is male sterile.These include use of manual or mechanical emasculation (or detasseling),cytoplasmic genetic male sterility, nuclear genetic male sterility,gametocides and the like.

Hybrid maize seed is often produced by a male sterility systemincorporating manual or mechanical detasseling. Alternate strips of twoinbred varieties of maize are planted in a field, and the pollen-bearingtassels are removed from one of the inbreds (female) prior to pollenshed. Providing that there is sufficient isolation from sources offoreign maize pollen, the ears of the detasseled inbred will befertilized only from the other inbred (male), and the resulting seed istherefore hybrid and will form hybrid plants.

The laborious detasseling process can be avoided by using cytoplasmicmale-sterile (CMS) inbreds. Plants of a CMS inbred are male sterile as aresult of factors resulting from the cytoplasmic, as opposed to thenuclear, genome. Thus, this characteristic is inherited exclusivelythrough the female parent in maize plants, since only the femaleprovides cytoplasm to the fertilized seed. CMS plants are fertilizedwith pollen from another inbred that is not male-sterile. Pollen fromthe second inbred may or may not contribute genes that make the hybridplants male-fertile. The same hybrid seed, a portion produced fromdetasseled fertile maize and a portion produced using the CMS system canbe blended to insure that adequate pollen loads are available forfertilization when the hybrid plants are grown.

There are several methods of conferring genetic male sterilityavailable, such as multiple mutant genes at separate locations withinthe genome that confer male sterility, as disclosed in U.S. Pat. Nos.4,654,465 and 4,727,219 to Brar et al. and chromosomal translocations asdescribed by Patterson in U.S. Pat. Nos. 3,861,709 and 3,710,511. Theseand all patents referred to are incorporated by reference. In additionto these methods, Albertsen et al., of Pioneer Hi-Bred, U.S. Pat. No.5,432,068, have developed a system of nuclear male sterility whichincludes: identifying a gene which is critical to male fertility;silencing this native gene which is critical to male fertility; removingthe native promoter from the essential male fertility gene and replacingit with an inducible promoter; inserting this genetically engineeredgene back into the plant; and thus creating a plant that is male sterilebecause the inducible promoter is not “on” resulting in the malefertility gene not being transcribed. Fertility is restored by inducing,or turning “on”, the promoter, which in turn allows the gene thatconfers male fertility to be transcribed.

There are many other methods of conferring genetic male sterility in theart, each with its own benefits and drawbacks. These methods use avariety of approaches such as delivering into the plant a gene encodinga cytotoxic substance associated with a male tissue specific promoter oran antisense system in which a gene critical to fertility is identifiedand an antisense to that gene is inserted in the plant (see:Fabinjanski, et al. EPO 89/3010153.8 Publication No. 329,308 and PCTApplication PCT/CA90/00037 published as WO 90/08828).

Another system useful in controlling male sterility makes use ofgametocides. Gametocides are not a genetic system, but rather a topicalapplication of chemicals. These chemicals affect cells that are criticalto male fertility. The application of these chemicals affects fertilityin the plants only for the growing season in which the gametocide isapplied (see Carlson, Glenn R., U.S. Pat. No. 4,936,904). Application ofthe gametocide, timing of the application and genotype specificity oftenlimit the usefulness of the approach and it is not appropriate in allsituations.

The use of male sterile inbreds is but one factor in the production ofmaize hybrids. The development of maize hybrids in a maize plantbreeding program requires, in general, the development of homozygousinbred lines, the crossing of these lines, and the evaluation of thecrosses. Maize plant breeding programs combine the genetic backgroundsfrom two or more inbred lines or various other germplasm sources intobreeding populations from which new inbred lines are developed byselfing and selection of desired phenotypes. Hybrids also can be used asa source of plant breeding material or as source populations from whichto develop or derive new maize lines. Plant breeding techniques known inthe art and used in a maize plant breeding program include, but are notlimited to, recurrent selection, backcrossing, double haploids, pedigreebreeding, restriction fragment length polymorphism enhanced selection,genetic marker enhanced selection, and transformation. Often acombination of these techniques are used. The inbred lines derived fromhybrids can be developed using plant breeding techniques as describedabove. New inbreds are crossed with other inbred lines and the hybridsfrom these crosses are evaluated to determine which of those havecommercial potential.

Backcrossing can be used to improve inbred lines and a hybrid which ismade using those inbreds. Backcrossing can be used to transfer aspecific desirable trait from one line, the donor parent, to an inbredcalled the recurrent parent which has overall good agronomiccharacteristics yet that lacks the desirable trait. This transfer of thedesirable trait into an inbred with overall good agronomiccharacteristics can be accomplished by first crossing a recurrent parentto a donor parent (non-recurrent parent). The progeny of this cross isthen mated back to the recurrent parent followed by selection in theresultant progeny for the desired trait to be transferred from thenon-recurrent parent. Typically after four or more backcross generationswith selection for the desired trait, the progeny will containessentially all genes of the recurrent parent except for the genescontrolling the desired trait. But the number of backcross generationscan be less if molecular markers are used during the selection or elitegermplasm is used as the donor parent. The last backcross generation isthen selfed to give pure breeding progeny for the gene(s) beingtransferred.

Backcrossing can also be used in conjunction with pedigree breeding todevelop new inbred lines. For example, an F1 can be created that isbackcrossed to one of its parent lines to create a BC1. Progeny areselfed and selected so that the newly developed inbred has many of theattributes of the recurrent parent and some of the desired attributes ofthe non-recurrent parent.

Recurrent selection is a method used in a plant breeding program toimprove a population of plants. The method entails individual plantscross pollinating with each other to form progeny which are then grown.The superior progeny are then selected by any number of methods, whichinclude individual plant, half sib progeny, full sib progeny, selfedprogeny and topcrossing. The selected progeny are cross pollinated witheach other to form progeny for another population. This population isplanted and again superior plants are selected to cross pollinate witheach other. Recurrent selection is a cyclical process and therefore canbe repeated as many times as desired. The objective of recurrentselection is to improve the traits of a population. The improvedpopulation can then be used as a source of breeding material to obtaininbred lines to be used in hybrids or used as parents for a syntheticcultivar. A synthetic cultivar is the resultant progeny formed by theintercrossing of several selected inbreds. Mass selection is a usefultechnique when used in conjunction with molecular marker enhancedselection.

Molecular markers including techniques such as Isozyme Electrophoresis,Restriction Fragment Length Polymorphisms (RFLPs), Randomly AmplifiedPolymorphic DNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction(AP-PCR), DNA Amplification Fingerprinting (DAF), Sequence CharacterizedAmplified Regions (SCARs), Amplified Fragment Length Polymorphisms(AFLPs), Single Nucleotide Polymorphisms (SNPs) and Simple SequenceRepeats (SSRs) may be used in plant breeding methods utilizing 33M72.One use of molecular markers is Quantitative Trait Loci (QTL) mapping.QTL mapping is the use of markers, which are closely linked to allelesthat have measurable effects on a quantitative trait. Selection in thebreeding process is based upon the accumulation of markers linked to thepositive effecting alleles and/or the elimination of the markers linkedto the negative effecting alleles from the plant's genome.

Molecular markers can also be used during the breeding process for theselection of qualitative traits. For example, markers closely linked toalleles or markers containing sequences within the actual alleles ofinterest can be used to select plants that contain the alleles ofinterest during a backcrossing breeding program. The markers can also beused to select for the genome of the recurrent parent and against themarkers of the donor parent. Using this procedure can minimize theamount of genome from the donor parent that remains in the selectedplants. It can also be used to reduce the number of crosses back to therecurrent parent needed in a backcrossing program. The use of molecularmarkers in the selection process is often called Genetic Marker EnhancedSelection.

The production of double haploids can also be used for the developmentof inbreds in a breeding program. Double haploids are produced by thedoubling of a set of chromosomes (1N) from a heterozygous plant toproduce a completely homozygous individual. For example, see Wan et al.,“Efficient Production of Doubled Haploid Plants Through ColchicineTreatment of Anther-Derived Maize Callus”, Theoretical and AppliedGenetics, 77:889–892, 1989 and U.S. Application 2003/0005479. This canbe advantageous because the process omits the generations of selfingneeded to obtain a homozygous plant from a heterozygous source.

Hybrid seed production requires elimination or inactivation of pollenproduced by the female parent. Incomplete removal or inactivation of thepollen provides the potential for self-pollination. This inadvertentlyself-pollinated seed may be unintentionally harvested and packaged withhybrid seed. Also, because the male parent is grown next to the femaleparent in the field there is the very low probability that the maleselfed seed could be unintentionally harvested and packaged with thehybrid seed. Once the seed from the hybrid bag is planted, it ispossible to identify and select these self-pollinated plants. Theseself-pollinated plants will be genetically equivalent to one of theinbred lines used to produce the hybrid. Though the possibility ofinbreds being included in hybrid seed bags exists, the occurrence isvery low because much care is taken to avoid such inclusions. It isworth noting that hybrid seed is sold to growers for the production ofgrain and forage and not for breeding or seed production.

By an individual skilled in plant breeding, these inbred plantsunintentionally included in commercial hybrid seed can be identified andselected due to their decreased vigor when compared to the hybrid.Inbreds are identified by their less vigorous appearance for vegetativeand/or reproductive characteristics, including shorter plant height,small ear size, ear and kernel shape, cob color, or othercharacteristics.

Identification of these self-pollinated lines can also be accomplishedthrough molecular marker analyses. See, “The Identification of FemaleSelfs in Hybrid Maize: A Comparison Using Electrophoresis andMorphology”, Smith, J. S. C. and Wych, R. D., Seed Science andTechnology 14, pp. 1–8 (1995), the disclosure of which is expresslyincorporated herein by reference. Through these technologies, thehomozygosity of the self pollinated line can be verified by analyzingallelic composition at various loci along the genome. Those methodsallow for rapid identification of the invention disclosed herein. Seealso, “Identification of Atypical Plants in Hybrid Maize Seed byPostcontrol and Electrophoresis” Sarca, V. et al., Probleme de GeneticaTeoritica si Aplicata Vol. 20 (1) pp. 29–42.

Another form of commercial hybrid production involves the use of amixture of male sterile hybrid seed and male pollinator seed. Whenplanted, the resulting male sterile hybrid plants are pollinated by thepollinator plants. This method is primarily used to produce grain withenhanced quality grain traits, such as high oil, because desired qualitygrain traits expressed in the pollinator will also be expressed in thegrain produced on the male sterile hybrid plant. In this method thedesired quality grain trait does not have to be incorporated by lengthyprocedures such as recurrent backcross selection into an inbred parentline. One use of this method is described in U.S. Pat. Nos. 5,704,160and 5,706,603.

There are many important factors to be considered in the art of plantbreeding, such as the ability to recognize important morphological andphysiological characteristics, the ability to design evaluationtechniques for genotypic and phenotypic traits of interest, and theability to search out and exploit the genes for the desired traits innew or improved combinations.

The objective of commercial maize hybrid line development resulting froma maize plant breeding program is to develop new inbred lines to producehybrids that combine to produce high grain yields and superior agronomicperformance. One of the primary traits breeders seek is yield. However,many other major agronomic traits are of importance in hybridcombination and have an impact on yield or otherwise provide superiorperformance in hybrid combinations. Such traits include percent grainmoisture at harvest, relative maturity, resistance to stalk breakage,resistance to root lodging, grain quality, and disease and insectresistance. In addition, the lines per se must have acceptableperformance for parental traits such as seed yields, kernel sizes,pollen production, all of which affect ability to provide parental linesin sufficient quantity and quality for hybridization. These traits havebeen shown to be under genetic control and many if not all of the traitsare affected by multiple genes.

A breeder uses various methods to help determine which plants should beselected from the segregating populations and ultimately which inbredlines will be used to develop hybrids for commercialization. In additionto the knowledge of the germplasm and other skills the breeder uses, apart of the selection process is dependent on experimental designcoupled with the use of statistical analysis. Experimental design andstatistical analysis are used to help determine which plants, whichfamily of plants, and finally which inbred lines and hybrid combinationsare significantly better or different for one or more traits ofinterest. Experimental design methods are used to assess error so thatdifferences between two inbred lines or two hybrid lines can be moreaccurately determined. Statistical analysis includes the calculation ofmean values, determination of the statistical significance of thesources of variation, and the calculation of the appropriate variancecomponents. Either a five or one percent significance level iscustomarily used to determine whether a difference that occurs for agiven trait is real or due to the environment or experimental error. Oneof ordinary skill in the art of plant breeding would know how toevaluate the traits of two plant varieties to determine if there is nosignificant difference between the two traits expressed by thosevarieties. For example, see Fehr, Walt, Principles of CultivarDevelopment, pp. 261–286 (1987) which is incorporated herein byreference. Mean trait values may be used to determine whether traitdifferences are significant, and preferably the traits are measured onplants grown under the same environmental conditions.

Combining ability of a line, as well as the performance of the line perse, is a factor in the selection of improved maize inbreds. Combiningability refers to a line's contribution as a parent when crossed withother lines to form hybrids. The hybrids formed for the purpose ofselecting superior lines are designated test crosses. One way ofmeasuring combining ability is by using breeding values. Breeding valuesare based in part on the overall mean of a number of test crosses. Thismean is then adjusted to remove environmental effects and it is adjustedfor known genetic relationships among the lines.

Once such a line is developed its value to society is substantial sinceit is important to advance the germplasm base as a whole in order tomaintain or improve traits such as yield, disease resistance, pestresistance and plant performance in extreme weather conditions.

SUMMARY OF THE INVENTION

According to the invention, there is provided a hybrid maize plant, andits parts designated as 33M72. This invention thus relates to the hybridseed 33M72, the hybrid plant and its parts produced from the seed, andvariants, mutants and trivial modifications of hybrid maize 33M72. Thisinvention also relates to methods for producing a maize plant containingin its genetic material one or more transgenes and to the transgenicmaize plants and their parts produced by those methods. This inventionfurther relates to methods for producing maize lines derived from hybridmaize 33M72 and to the maize lines derived by the use of those methods.This hybrid maize plant is characterized by good yield and good overallagronomics.

Definitions

Certain definitions used in the specification are provided below. Inorder to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided. NOTE: ABS is in absolute termsand % MN is percent of the mean for the experiments in which the inbredor hybrid was grown. PCT designates that the trait is calculated as apercentage. % NOT designates the percentage of plants that did notexhibit trait. For example, STKLDG % NOT is the percentage of plants ina plot that were not stalk lodged. These designators will follow thedescriptors to denote how the values are to be interpreted. Below arethe descriptors used in the data tables included herein.

ABTSTK=ARTIFICIAL BRITTLE STALK. A count of the number of “snapped”plants per plot following machine snapping. A snapped plant has itsstalk completely snapped at a node between the base of the plant and thenode above the ear. Expressed as percent of plants that did not snap.

ADF=PERCENT ACID DETERGENT FIBER. The percent of dry matter that is aciddetergent fiber in chopped whole plant forage.

ALLELE. Any of one or more alternative forms of a genetic sequence. In adiploid cell or organism, the two alleles of a given sequence typicallyoccupy corresponding loci on a pair of homologous chromosomes.

ANTROT=ANTHRACNOSE STALK ROT (Colletotrichum graminicola). A 1 to 9visual rating indicating the resistance to Anthracnose Stalk Rot. Ahigher score indicates a higher resistance.

BACKCROSSING. Process in which a breeder crosses a hybrid progeny lineback to one of the parental genotypes one or more times.

BARPLT=BARREN PLANTS. The percent of plants per plot that were notbarren (lack ears).

BREEDING. The genetic manipulation of living organisms.

BREEDING CROSS. A cross to introduce new genetic material into a plantfor the development of a new variety. For example, one could cross plantA with plant B, wherein plant B would be genetically different fromplant A. After the breeding cross, the resulting F1 plants could then beselfed or sibbed for one, two, three or more times (F1, F2, F3, etc.)until a new inbred variety is developed. For clarification, such newinbred varieties would be within a pedigree distance of one breedingcross of plants A and B. The process described above would be referredto as one breeding cycle.

BRTSTK=BRITTLE STALKS. This is a measure of the stalk breakage near thetime of pollination, and is an indication of whether a hybrid or inbredwould snap or break near the time of flowering under severe winds. Dataare presented as percentage of plants that did not snap.

CELL. Cell as used herein includes a plant cell, whether isolated, intissue culture or incorporated in a plant or plant part.

CLDTST=COLD TEST. The percent of plants that germinate under cold testconditions.

CLN=CORN LETHAL NECROSIS. Synergistic interaction of maize chloroticmottle virus (MCMV) in combination with either maize dwarf mosaic virus(MDMV-A or MDMV-B) or wheat streak mosaic virus (WSMV). A 1 to 9 visualrating indicating the resistance to Corn Lethal Necrosis. A higher scoreindicates a higher resistance.

COMRST=COMMON RUST (Puccinia sorghi). A 1 to 9 visual rating indicatingthe resistance to Common Rust. A higher score indicates a higherresistance.

CP=PERCENT OF CRUDE PROTEIN. The percent of dry matter that is crudeprotein in chopped whole plant forage.

CROSS POLLINATION. A plant is cross pollinated if the pollen comes froma flower on a different plant from a different family or line. Crosspollination excludes sib and self pollination.

CROSS. As used herein, the term “cross” or “crossing” can refer to asimple X by Y cross, or the process of backcrossing, depending on thecontext.

D/D=DRYDOWN. This represents the relative rate at which a hybrid willreach acceptable harvest moisture compared to other hybrids on a 1 to 9rating scale. A high score indicates a hybrid that dries relatively fastwhile a low score indicates a hybrid that dries slowly.

DIPERS=DIPLODIA EAR MOLD SCORES (Diplodia maydis and Diplodiamacrospora). A 1 to 9 visual rating indicating the resistance toDiplodia Ear Mold. A higher score indicates a higher resistance.

DIPLOID PLANT PART. Refers to a plant part or cell that has the samediploid genotype as 33M72.

DIPROT=DIPLODIA STALK ROT SCORE. Score of stalk rot severity due toDiplodia (Diplodia maydis). Expressed as a 1 to 9 score with 9 beinghighly resistant.

DM=PERCENT OF DRY MATTER. The percent of dry material in chopped wholeplant silage.

DRPEAR=DROPPED EARS. A measure of the number of dropped ears per plotand represents the percentage of plants that did not drop ears prior toharvest.

D/T=DROUGHT TOLERANCE. This represents a 1 to 9 rating for droughttolerance, and is based on data obtained under stress conditions. A highscore indicates good drought tolerance and a low score indicates poordrought tolerance.

EARHT=EAR HEIGHT. The ear height is a measure from the ground to thehighest placed developed ear node attachment and is measured incentimeters.

EARMLD=GENERAL EAR MOLD. Visual rating (1 to 9 score) where a “1” isvery susceptible and a “9” is very resistant. This is based on overallrating for ear mold of mature ears without determining the specific moldorganism, and may not be predictive for a specific ear mold.

EARSZ=EAR SIZE. A 1 to 9 visual rating of ear size. The higher therating the larger the ear size.

EBTSTK=EARLY BRITTLE STALK. A count of the number of “snapped” plantsper plot following severe winds when the corn plant is experiencing veryrapid vegetative growth in the V5–V8 stage. Expressed as percent ofplants that did not snap.

ECB1LF=EUROPEAN CORN BORER FIRST GENERATION LEAF FEEDING (Ostrinianubilalis). A 1 to 9 visual rating indicating the resistance topreflowering leaf feeding by first generation European Corn Borer. Ahigher score indicates a higher resistance.

ECB2IT=EUROPEAN CORN BORER SECOND GENERATION INCHES OF TUNNELING(Ostrinia nubilalis). Average inches of tunneling per plant in thestalk.

ECB2SC=EUROPEAN CORN BORER SECOND GENERATION (Ostrinia nubilalis). A 1to 9 visual rating indicating post flowering degree of stalk breakageand other evidence of feeding by European Corn Borer, Second Generation.A higher score indicates a higher resistance.

ECBDPE=EUROPEAN CORN BORER DROPPED EARS (Ostrinia nubilalis). Droppedears due to European Corn Borer. Percentage of plants that did not dropears under second generation corn borer infestation.

EGRWTH=EARLY GROWTH. This is a measure of the relative height and sizeof a corn seedling at the 2–4 leaf stage of growth. This is a visualrating (1 to 9), with 1 being weak or slow growth, 5 being averagegrowth and 9 being strong growth. Taller plants, wider leaves, moregreen mass and darker color constitute a higher score.

ELITE INBRED. An inbred that contributed desirable qualities when usedto produce commercial hybrids. An elite inbred may also be used infurther breeding for the purpose of developing further improvedvarieties.

ERTLDG=EARLY ROOT LODGING. Early root lodging is the percentage ofplants that do not root lodge prior to or around anthesis; plants thatlean from the vertical axis at an approximately 30 degree angle orgreater would be counted as root lodged.

ERTLPN=EARLY ROOT LODGING. An estimate of the percentage of plants thatdo not root lodge prior to or around anthesis; plants that lean from thevertical axis at an approximately 30 degree angle or greater would beconsidered as root lodged.

ERTLSC=EARLY ROOT LODGING SCORE. Score for severity of plants that leanfrom a vertical axis at an approximate 30 degree angle or greater whichtypically results from strong winds prior to or around floweringrecorded within 2 weeks of a wind event. Expressed as a 1 to 9 scorewith 9 being no lodging.

ESTCNT=EARLY STAND COUNT. This is a measure of the stand establishmentin the spring and represents the number of plants that emerge on perplot basis for the inbred or hybrid.

EYESPT=EYE SPOT (Kabatiella zeae or Aureobasidium zeae). A 1 to 9 visualrating indicating the resistance to Eye Spot. A higher score indicates ahigher resistance.

FUSERS=FUSARIUM EAR ROT SCORE (Fusarium moniliforme or Fusariumsubglutinans). A 1 to 9 visual rating indicating the resistance toFusarium ear rot. A higher score indicates a higher resistance.

GDU=Growing Degree Units. Using the Barger Heat Unit Theory, whichassumes that maize growth occurs in the temperature range 50° F.–86° F.and that temperatures outside this range slow down growth; the maximumdaily heat unit accumulation is 36 and the minimum daily heat unitaccumulation is 0. The seasonal accumulation of GDU is a major factor indetermining maturity zones.

GDUSHD=GDU TO SHED. The number of growing degree units (GDUs) or heatunits required for an inbred line or hybrid to have approximately 50percent of the plants shedding pollen and is measured from the time ofplanting. Growing degree units are calculated by the Barger Method,where the heat units for a 24-hour period are:

${GDU} = {\frac{( {{Max}.\mspace{14mu}{temp}.{+ {{Min}.\mspace{14mu}{temp}.}}} )}{2} - 50}$

The highest maximum temperature used is 86° F. and the lowest minimumtemperature used is 50° F. For each inbred or hybrid it takes a certainnumber of GDUs to reach various stages of plant development.

GDUSLK=GDU TO SILK. The number of growing degree units required for aninbred line or hybrid to have approximately 50 percent of the plantswith silk emergence from time of planting. Growing degree units arecalculated by the Barger Method as given in GDU SHD definition.

GENOTYPE. Refers to the genetic constitution of a cell or organism.

GIBERS=GIBBERELLA EAR ROT (PINK MOLD) (Gibberella zeae). A 1 to 9 visualrating indicating the resistance to Gibberella Ear Rot. A higher scoreindicates a higher resistance.

GIBROT=GIBBERELLA STALK ROT SCORE. Score of stalk rot severity due toGibberella (Gibberella zeae). Expressed as a 1 to 9 score with 9 beinghighly resistant.

GLFSPT=GRAY LEAF SPOT (Cercospora zeae-maydis). A 1 to 9 visual ratingindicating the resistance to Gray Leaf Spot. A higher score indicates ahigher resistance.

GOSWLT=GOSS' WILT (Corynebacterium nebraskense). A 1 to 9 visual ratingindicating the resistance to Goss' Wilt. A higher score indicates ahigher resistance.

GRNAPP=GRAIN APPEARANCE. This is a 1 to 9 rating for the generalappearance of the shelled grain as it is harvested based on such factorsas the color of harvested grain, any mold on the grain, and any crackedgrain. High scores indicate good grain quality.

H/POP=YIELD AT HIGH DENSITY. Yield ability at relatively high plantdensities on a 1 to 9 relative rating system with a higher numberindicating the hybrid responds well to high plant densities for yieldrelative to other hybrids. A 1, 5, and 9 would represent very poor,average, and very good yield response, respectively, to increased plantdensity.

HCBLT=HELMINTHOSPORIUM CARBONUM LEAF BLIGHT (Helminthosporium carbonum).A 1 to 9 visual rating indicating the resistance to Helminthosporiuminfection. A higher score indicates a higher resistance.

HD SMT=HEAD SMUT (Sphacelotheca reiliana). This score indicates thepercentage of plants not infected.

HSKCVR=HUSK COVER. A 1 to 9 score based on performance relative to keychecks, with a score of 1 indicating very short husks, tip of ear andkernels showing; 5 is intermediate coverage of the ear under mostconditions, sometimes with thin husk; and a 9 has husks extending andclosed beyond the tip of the ear. Scoring can best be done nearphysiological maturity stage or any time during dry down untilharvested.

INC D/A=GROSS INCOME (DOLLARS PER ACRE). Relative income per acreassuming drying costs of two cents per point above 15.5 percent harvestmoisture and current market price per bushel.

INCOME/ACRE. Income advantage of hybrid to be patented over other hybridon per acre basis.

INC ADV=GROSS INCOME ADVANTAGE. GROSS INCOME advantage of variety #1over variety #2.

KSZDCD=KERNEL SIZE DISCARD. The percent of discard seed; calculated asthe sum of discarded tip kernels and extra large kernels.

LINKAGE. Refers to a phenomenon wherein alleles on the same chromosometend to segregate together more often than expected by chance if theirtransmission was independent.

LINKAGE DISEQUILIBRIUM. Refers to a phenomenon wherein alleles tend toremain together in linkage groups when segregating from parents tooffspring, with a greater frequency than expected from their individualfrequencies.

L/POP=YIELD AT LOW DENSITY. Yield ability at relatively low plantdensities on a 1 to 9 relative system with a higher number indicatingthe hybrid responds well to low plant densities for yield relative toother hybrids. A 1, 5, and 9 would represent very poor, average, andvery good yield response, respectively, to low plant density.

LRTLDG=LATE ROOT LODGING. Late root lodging is the percentage of plantsthat do not root lodge after anthesis through harvest; plants that leanfrom the vertical axis at an approximately 30 degree angle or greaterwould be counted as root lodged.

LRTLPN=LATE ROOT LODGING. Late root lodging is an estimate of thepercentage of plants that do not root lodge after anthesis throughharvest; plants that lean from the vertical axis at an approximately 30degree angle or greater would be considered as root lodged.

LRTLSC=LATE ROOT LODGING SCORE. Score for severity of plants that leanfrom a vertical axis at an approximate 30 degree angle or greater whichtypically results from strong winds after flowering. Recorded prior toharvest when a root-lodging event has occurred. This lodging results inplants that are leaned or “lodged” over at the base of the plant and donot straighten or “goose-neck” back to a vertical position. Expressed asa 1 to 9 score with 9 being no lodging.

MDMCPX=MAIZE DWARF MOSAIC COMPLEX (MDMV=Maize Dwarf Mosaic Virus andMCDV=Maize Chlorotic Dwarf Virus). A 1 to 9 visual rating indicating theresistance to Maize Dwarf Mosaic Complex. A higher score indicates ahigher resistance.

MST=HARVEST MOISTURE. The moisture is the actual percentage moisture ofthe grain at harvest.

MSTADV=MOISTURE ADVANTAGE. The moisture advantage of variety #1 overvariety #2 as calculated by: MOISTURE of variety #2−MOISTURE of variety#1=MOISTURE ADVANTAGE of variety #1.

NLFBLT=NORTHERN LEAF BLIGHT (Helminthosporium turcicum or Exserohilumturcicum). A 1 to 9 visual rating indicating the resistance to NorthernLeaf Blight. A higher score indicates a higher resistance.

OILT=GRAIN OIL. Absolute value of oil content of the kernel as predictedby Near-Infrared Transmittance and expressed as a percent of dry matter.

PEDIGREE DISTANCE. Relationship among generations based on theirancestral links as evidenced in pedigrees. May be measured by thedistance of the pedigree from a given starting point in the ancestry.

PERCENT IDENTITY. Percent identity as used herein refers to thecomparison of the alleles of two plants or lines as scored by matchingloci. Percent identity is determined by comparing a statisticallysignificant number of the loci of two plants or lines and scoring amatch when the same two alleles are present at the same loci for eachplant. For example, a percent identity of 90% between hybrid 33M72 andanother plant means that the two plants have the same two alleles at 90%of their loci.

PERCENT SIMILARITY. Percent similarity as used herein refers to thecomparison of the alleles of two plants or lines as scored by matchingalleles. Percent similarity is determined by comparing a statisticallysignificant number of the loci of two plants or lines and scoring oneallele match when the same allele is present at the same loci for eachplant and two allele matches when the same two alleles are present atthe same loci for each plant. A percent similarity of 90% between hybrid33M72 and another plant means that the two plants have 90% matchingalleles.

PLANT. As used herein, the term “plant” includes reference to animmature or mature whole plant, including a plant that has beendetasseled or from which seed or grain has been removed. Seed or embryothat will produce the plant is also considered to be the plant.

PLANT PARTS. As used herein, the term “plant parts” includes leaves,stems, roots, seed, grain, embryo, pollen, ovules, flowers, ears, cobs,husks, stalks, root tips, anthers, pericarp, silk, tissue, cells and thelike.

PLTHT=PLANT HEIGHT. This is a measure of the height of the plant fromthe ground to the tip of the tassel in centimeters.

POLSC=POLLEN SCORE. A 1 to 9 visual rating indicating the amount ofpollen shed. The higher the score the more pollen shed.

POLWT=POLLEN WEIGHT. This is calculated by dry weight of tasselscollected as shedding commences minus dry weight from similar tasselsharvested after shedding is complete.

POP K/A=PLANT POPULATIONS. Measured as 1000 s per acre.

POP ADV=PLANT POPULATION ADVANTAGE. The plant population advantage ofvariety #1 over variety #2 as calculated by PLANT POPULATION of variety#2−PLANT POPULATION of variety #1=PLANT POPULATION ADVANTAGE of variety#1.

PRM=PREDICTED RELATIVE MATURITY. This trait, predicted relativematurity, is based on the harvest moisture of the grain. The relativematurity rating is based on a known set of checks and utilizes standardlinear regression analyses and is also referred to as the ComparativeRelative Maturity Rating System that is similar to the MinnesotaRelative Maturity Rating System.

PRMSHD=A relative measure of the growing degree units (GDU) required toreach 50% pollen shed. Relative values are predicted values from thelinear regression of observed GDU's on relative maturity of commercialchecks.

PROT=GRAIN PROTEIN. Absolute value of protein content of the kernel aspredicted by Near-infrared Transmittance and expressed as a percent ofdry matter.

RTLDG=ROOT LODGING. Root lodging is the percentage of plants that do notroot lodge; plants that lean from the vertical axis at an approximately30 degree angle or greater would be counted as root lodged.

RTLADV=ROOT LODGING ADVANTAGE. The root lodging advantage of variety #1over variety #2.

SCTGRN=SCATTER GRAIN. A 1 to 9 visual rating indicating the amount ofscatter grain (lack of pollination or kernel abortion) on the ear. Thehigher the score the less scatter grain.

SDGVGR=SEEDLING VIGOR. This is the visual rating (1 to 9) of the amountof vegetative growth after emergence at the seedling stage(approximately five leaves). A higher score indicates better vigor.

SEL IND=SELECTION INDEX. The selection index gives a single measure ofthe hybrid's worth based on information for up to five traits. A maizebreeder may utilize his or her own set of traits for the selectionindex. One of the traits that is almost always included is yield. Theselection index data presented in the tables represent the mean valueaveraged across testing stations.

SIL DMP=SILAGE DRY MATTER. The percent of dry material in chopped wholeplant silage.

SELF POLLINATION. A plant is self-pollinated if pollen from one floweris transferred to the same or another flower of the same plant.

SIB POLLINATION. A plant is sib-pollinated when individuals within thesame family or line are used for pollination.

SLFBLT=SOUTHERN LEAF BLIGHT (Helminthosporium maydis or Bipolarismaydis). A 1 to 9 visual rating indicating the resistance to SouthernLeaf Blight. A higher score indicates a higher resistance.

SOURST=SOUTHERN RUST (Puccinia polysora). A 1 to 9 visual ratingindicating the resistance to Southern Rust. A higher score indicates ahigher resistance.

STAGRN=STAY GREEN. Stay green is the measure of plant health near thetime of black layer formation (physiological maturity). A high scoreindicates better late-season plant health.

STARCH=PERCENT OF STARCH. The percent of dry matter that is starch inchopped whole plant forage.

STDADV=STALK STANDING ADVANTAGE. The advantage of variety #1 overvariety #2 for the trait STK CNT.

STKCNT=NUMBER OF PLANTS. This is the final stand or number of plants perplot.

STKLDG=STALK LODGING REGULAR. This is the percentage of plants that didnot stalk lodge (stalk breakage) at regular harvest (when grain moistureis between about 20 and 30%) as measured by either natural lodging orpushing the stalks and determining the percentage of plants that breakbelow the ear.

STKLDL=LATE STALK LODGING. This is the percentage of plants that did notstalk lodge (stalk breakage) at or around late season harvest (whengrain moisture is between about 15 and 18%) as measured by eithernatural lodging or pushing the stalks and determining the percentage ofplants that break below the ear.

STKLDS=STALK LODGING SCORE. A plant is considered as stalk lodged if thestalk is broken or crimped between the ear and the ground. This can becaused by any or a combination of the following: strong winds late inthe season, disease pressure within the stalks, ECB damage orgenetically weak stalks. This trait should be taken just prior to or atharvest. Expressed on a 1 to 9 scale with 9 being no lodging.

STLLPN=LATE STALK LODGING. This is the percent of plants that did notstalk lodge (stalk breakage or crimping) at or around late seasonharvest (when grain moisture is below 20%) as measured by either naturallodging or pushing the stalks and determining the percentage of plantsthat break or crimp below the ear.

STLPCN=STALK LODGING REGULAR. This is an estimate of the percentage ofplants that did not stalk lodge (stalk breakage at regular harvest (whengrain moisture is between about 20 and 30%) as measured by eithernatural lodging or pushing the stalks and determining the percentage ofplants that break below the ear.

STRT=GRAIN STARCH. Absolute value of starch content of the kernel aspredicted by Near-Infrared Transmittance and expressed as a percent ofdry matter.

STWWLT=Stewart's Wilt (Erwinia stewartii). A 1 to 9 visual ratingindicating the resistance to Stewart's Wilt. A higher score indicates ahigher resistance.

TASBLS=TASSEL BLAST. A 1 to 9 visual rating was used to measure thedegree of blasting (necrosis due to heat stress) of the tassel at thetime of flowering. A 1 would indicate a very high level of blasting attime of flowering, while a 9 would have no tassel blasting.

TASSZ=TASSEL SIZE. A 1 to 9 visual rating was used to indicate therelative size of the tassel. The higher the rating the larger thetassel.

TAS WT=TASSEL WEIGHT. This is the average weight of a tassel (grams)just prior to pollen shed.

TDM/HA=TOTAL DRY MATTER PER HECTARE. Yield of total dry plant materialin metric tons per hectare.

TEXEAR=EAR TEXTURE. A 1 to 9 visual rating was used to indicate therelative hardness (smoothness of crown) of mature grain. A 1 would bevery soft (extreme dent) while a 9 would be very hard (flinty or verysmooth crown).

TILLER=TILLERS. A count of the number of tillers per plot that couldpossibly shed pollen was taken. Data are given as a percentage oftillers: number of tillers per plot divided by number of plants perplot.

TST WT=TEST WEIGHT (UNADJUSTED). The measure of the weight of the grainin pounds for a given volume (bushel).

TSWADV=TEST WEIGHT ADVANTAGE. The test weight advantage of variety #1over variety #2.

WIN M %=PERCENT MOISTURE WINS.

WIN Y %=PERCENT YIELD WINS.

YIELD=YIELD OF SILAGE. Yield in tons per acre at 30% dry matter.

YIELD BU/A=YIELD (BUSHELS/ACRE). Yield of the grain at harvest inbushels per acre adjusted to 15% moisture.

YLDADV=YIELD ADVANTAGE. The yield advantage of variety #1 over variety#2 as calculated by: YIELD of variety #1−YIELD variety #2=yieldadvantage of variety #1.

YLD SC=YIELD SCORE. A 1 to 9 visual rating was used to give a relativerating for yield based on plot ear piles. The higher the rating thegreater visual yield appearance.

Definitions for Area of Adaptability

When referring to area of adaptability, such term is used to describethe location with the environmental conditions that would be well suitedfor this maize line. Area of adaptability is based on a number offactors, for example: days to maturity, insect resistance, diseaseresistance, and drought resistance. Area of adaptability does notindicate that the maize line will grow in every location within the areaof adaptability or that it will not grow outside the area.

-   Central Corn Belt: Iowa, Illinois, Indiana-   Drylands: non-irrigated areas of North Dakota, South Dakota,    Nebraska, Kansas, Colorado and Oklahoma-   Eastern U.S.: Ohio, Pennsylvania, Delaware, Maryland, Virginia, and    West Virginia-   North central U.S.: Minnesota and Wisconsin-   Northeast: Michigan, New York, Vermont, and Ontario and Quebec    Canada-   Northwest U.S.: North Dakota, South Dakota, Wyoming, Washington,    Oregon, Montana, Utah, and Idaho-   South central U.S.: Missouri, Tennessee, Kentucky, Arkansas-   Southeast U.S.: North Carolina, South Carolina, Georgia, Florida,    Alabama, Mississippi, and Louisiana-   Southwest U.S.: Texas, Oklahoma, New Mexico, Arizona-   Western U.S.: Nebraska, Kansas, Colorado, and California-   Maritime Europe: Northern France, Germany, Belgium, Netherlands and    Austria

DETAILED DESCRIPTION OF THE INVENTION

Inbred maize lines are typically developed for use in the production ofhybrid maize lines. Maize hybrids need to be highly homogeneous,heterozygous and reproducible to be useful as commercial hybrids. Thereare many analytical methods available to determine the heterozygousnature and the identity of these lines.

The oldest and most traditional method of analysis is the observation ofphenotypic traits. The data is usually collected in field experimentsover the life of the maize plants to be examined. Phenotypiccharacteristics most often observed are for traits associated with plantmorphology, ear and kernel morphology, insect and disease resistance,maturity, and yield.

In addition to phenotypic observations, the genotype of a plant can alsobe examined. A plant's genotype can be used to identify plants of thesame variety or a related variety. For example, the genotype can be usedto determine the pedigree of a plant. There are many laboratory-basedtechniques available for the analysis, comparison and characterizationof plant genotype; among these are Isozyme Electrophoresis, RestrictionFragment Length Polymorphisms (RFLPs), Randomly Amplified PolymorphicDNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction (AP-PCR), DNAAmplification Fingerprinting (DAF), Sequence Characterized AmplifiedRegions (SCARs), Amplified Fragment Length Polymorphisms (AFLPs), SimpleSequence Repeats (SSRs) which are also referred to as Microsatellites,and Single Nucleotide Polymorphisms (SNPs).

Isozyme Electrophoresis and RFLPs as discussed in Lee, M., “Inbred Linesof Maize and Their Molecular Markers,” The Maize Handbook,(Springer-Verlag, New York, Inc. 1994, at 423–432) incorporated hereinby reference, have been widely used to determine genetic composition.Isozyme Electrophoresis has a relatively low number of available markersand a low number of allelic variants. RFLPs allow more discriminationbecause they have a higher degree of allelic variation in maize and alarger number of markers can be found. Both of these methods have beeneclipsed by SSRs as discussed in Smith et al., “An evaluation of theutility of SSR loci as molecular markers in maize (Zea mays L.):comparisons with data from RFLPs and pedigree”, Theoretical and AppliedGenetics (1997) vol. 95 at 163–173 and by Pejic et al., “Comparativeanalysis of genetic similarity among maize inbreds detected by RFLPs,RAPDs, SSRs, and AFLPs,” Theoretical and Applied Genetics (1998) at1248–1255 incorporated herein by reference. SSR technology is moreefficient and practical to use than RFLPs; more marker loci can beroutinely used and more alleles per marker locus can be found using SSRsin comparison to RFLPs. Single Nucleotide Polymorphisms may also be usedto identify the unique genetic composition of the invention and progenylines retaining that unique genetic composition. Various molecularmarker techniques may be used in combination to enhance overallresolution.

Maize DNA molecular marker linkage maps have been rapidly constructedand widely implemented in genetic studies. One such study is describedin Boppenmaier, et al., “Comparisons among strains of inbreds forRFLPs”, Maize Genetics Cooperative Newsletter, 65:1991, pg. 90, isincorporated herein by reference.

Pioneer Brand Hybrid 33M72 is characterized by good yield and goodoverall agronomics. Hybrid 33M72 further demonstrates good stalkstrength, good late season root strength, and good stay green. Hybrid33M72 exhibits a substantial degree of Bacillus thuringiensis resistanceto Corn Rootworm, excellent Goss' Wilt tolerance, good tolerance toNorthern Leaf Blight, Diplodia Ear Rot tolerance and good tolerance toFusarium Ear and Kernel Rot. The hybrid is broadly adapted and isparticularly suited to the Central Corn Belt, Eastern, and Western areasof the United States.

Pioneer Brand Hybrid 33M72 is a single cross, yellow endosperm, dentmaize hybrid. Hybrid 33M72 has a relative maturity of approximately 112based on the Comparative Relative Maturity Rating System for harvestmoisture of grain.

This hybrid has the following characteristics based on the datacollected primarily at Johnston, Iowa.

TABLE 1 VARIETY DESCRIPTION INFORMATION 33M72 AVG STDEV N 1. TYPE:(Describe intermediate types in comments section) 1 = Sweet, 2 = Dent, 3= Flint, 4 = Flour, 5 = Pop and  2 6 = Ornamental. Comments: Dent-Flint2. MATURITY: DAYS HEAT UNITS Days H. Units Emergence to 50% of plants insilk 61 1,257 Emergence to 50% of plants in pollen shed 61 1,257 10% to90% pollen shed  2   48 50% Silk to harvest at 25% moisture 3. PLANT:Plant Height (to tassel tip) (cm) 316.9 15.94 15 Ear Height (to base oftop ear node) (cm) 132.3 13.33 15 Length of Top Ear Internode (cm) 21.11.28 15 Average Number of Tillers per Plant 0.0 0.01 3 Average Number ofEars per Stalk 1.0 0.03 3 Anthocyanin of Brace Roots: 1 = Absent, 2 =Faint, 1 3 = Moderate, 4 = Dark 4. LEAF: Width of Ear Node Leaf (cm) 9.90.70 15 Length of Ear Node Leaf (cm) 98.1 3.63 15 Number of Leaves aboveTop Ear 6.7 0.82 15 Leaf Angle: (at anthesis, 2nd leaf above ear to 28.35.35 15 stalk above leaf) (Degrees) *Leaf Color: V. Dark Green Munsell:5GY36 Leaf Sheath Pubescence: 1 = none to 9 = like peach fuzz 5 5.TASSEL: Number of Primary Lateral Branches 11.7 1.50 15 Branch Anglefrom Central Spike 31.3 5.51 15 Tassel Length: (from peduncle node totassel tip), (cm). 64.9 6.37 15 Pollen Shed: 0 = male sterile, 9 = heavyshed 8 *Anther Color: Pale Yellow Munsell: 5Y86 *Glume Color: LightGreen Munsell: 5GY76 *Bar Glumes (glume bands): 1 = absent, 2 = present1 Peduncle Length: (from top leaf node to lower florets or 28.1 3.35 15branches), (cm). 6a. EAR (Unhusked ear) *Silk color: Red Munsell: 10RP38(3 days after silk emergence) *Fresh husk color: Med. Green Munsell:5GY68 *Dry husk color: White Munsell: 2.5Y92 (65 days after 50% silking)Ear position at dry husk stage: 1 = upright, 2 = horizontal, 1 3 =pendant Husk Tightness: (1 = very loose, 9 = very tight) 6 HuskExtension (at harvest): 1 = short(ears exposed), 2 2 = medium (<8 cm), 3= long (8–10 cm), 4 = v. long (>10 cm) 6b. EAR (Husked ear data) EarLength (cm): 17.4 1.06 15 Ear Diameter at mid-point (mm) 49.8 1.47 15Ear Weight (gm): 215.2 23.88 15 Number of Kernel Rows: 17.7 1.67 15Kernel Rows: 1 = indistinct, 2 = distinct 2 Row Alignment: 1 = straight,2 = slightly curved, 3 = spiral 2 Shank Length (cm): 10.6 1.35 15 EarTaper: 1 = slight cylind., 2 = average, 3 = extreme 2 7. KERNEL (Dried):Kernel Length (mm): 14.3 0.62 15 Kernel Width (mm): 7.4 0.51 15 KernelThickness (mm): 4.3 0.49 15 Round Kernels (shape grade) (%) 14.3 3.61 3Aleurone Color Pattern: 1 = homozygous, 2 = segregating 1 *AleuroneColor: Yellow Munsell: 10YR714 *Hard Endo. Color: Yellow Munsell:10YR712 Endosperm Type: 3 1 = sweet (su1), 2 = extra sweet (sh2), 3 =normal starch, 4 = high amylose starch, 5 = waxy starch, 6 = highprotein, 7 = high lysine, 8 = super sweet (se), 9 = high oil, 10 = otherWeight per 100 Kernels (unsized sample) (gm): 36.3 1.15 3 8. COB: *CobDiameter at mid-point (mm): 24.6 1.06 15 *Cob Color: Pink Munsell: 10R6610. DISEASE RESISTANCE: (Rate from 1 = most-susceptable to 9 =most-resistant. Leave blank if not tested, leave race or strain optionsblank if polygenic.) A. LEAF BLIGHTS, WILTS, AND LOCAL INFECTIONDISEASES Anthracnose Leaf Blight (Colletotrichum graminicola) CommonRust (Puccinia sorghi) Common Smut (Ustilago maydis) Eyespot (Kabatiellazeae) 8 Gross's Wilt (Clavibacter michiganense spp. nebraskense) 4 GrayLeaf Spot (Cercospora zeae-maydis) Helminthosporium Leaf Spot (Bipolariszeicola) Race: 6 Northern Leaf Blight (Exserohilum turcicum) Race: 3Southern Leaf Blight (Bipolaris maydis) Race: Southern Rust (Pucciniapolysora) Stewart's Wilt (Erwinia stewartii) Other (Specify):           B. SYSTEMIC DISEASES Corn Lethal Necrosis (MCMV and MDMV) Head Smut(Sphacelotheca reiliana) Maize Chlorotic Dwarf Virus (MDV) MaizeChlorotic Mottle Virus (MCMV) Maize Dwarf Mosaic Virus (MDMV) SorghumDowny Mildew of Corn (Peronosclerospora sorghi) Other (Specify):           C. STALK ROTS 4 Anthracnose Stalk Rot (Colletotrichumgraminicola) Diplodia Stalk Rot (Stenocarpella maydis) Fusarium StalkRot (Fusarium moniliforme) Gibberella Stalk Rot (Gibberella zeae) Other(Specify):            D. EAR AND KERNEL ROTS Aspergillus Ear and KernelRot (Aspergillus flavus) 5 Diplodia Ear Rot (Stenocarpella maydis) 6Fusarium Ear and Kernel Rot (Fusarium moniliforme) Gibberella Ear Rot(Gibberella zeae) Other (Specify):            11. INSECT RESISTANCE:(Rate from 1 = most-suscept. to 9 = most-resist., leave blank if nottested.) Corn Worm (Helicoverpa zea)     Leaf Feeding     Silk Feeding    Ear Damage Corn Leaf Aphid (Rophalosiphum maydis) Corn Sap Beetle(Capophilus dimidiatus) European Corn Borer (Ostrinia nubilalis) 1st.Generation (Typically whorl leaf feeding) 2nd. Generation (Typicallyleaf sheath-collar feeding)     Stalk Tunneling     cm tunneled/plantFall armyworm (Spodoptera fruqiperda)     Leaf Feeding     Silk Feeding    mg larval wt. Maize Weevil (Sitophilus zeamaize) Northern Rootworm(Diabrotica barberi) Southern Rootworm (Diabrotica undecimpunctata)Southwestern Corn Borer (Diatreaea grandiosella)     Leaf Feeding    Stalk Tunneling     cm tunneled/plant Two-spotted Spider Mite(Tetranychus utricae) Western Rootworm (Diabrotica virgifrea virgifrea)Other (Specify):            12. AGRONOMIC TRAITS: 4 Staygreen (at 65days after anthesis; rate from 1-worst to 9-excellent) % Dropped Ears(at 65 days after anthesis) % Pre-anthesis Brittle Snapping %Pre-anthesis Root Lodging % Post-anthesis Root Lodging (at 65 days afteranthesis) % Post-anthesis Stalk Lodging 12,058.0 Kg/ha (Yield at 12–13%grain moisture) *Munsell Glossy Book of Color, (A standard colorreference). Kollmorgen Inst. Corp. New Windsor, NY.

Research Comparisons for Pioneer Hybrid 33M72

Comparisons of characteristics for Pioneer Brand Hybrid 33M72 were madewith Hybrid 34H31, 33P66, 34N43 and 33M54.

Table 2A compares Pioneer Brand Hybrid 33M72 and Hybrid 34H31, a closelyrelated hybrid. As shown in the table, significant differences betweenHybrid 33M72 and Hybrid 34H31 are reported. It is expected that othersignificant differences would be observed if additional traits wereexamined.

Table 2B compares Pioneer Brand Hybrid 33M72 and Hybrid 33P66, asimilarly adapted hybrid. Significant differences between Hybrid 33M72and Hybrid 33P66 include early growth score, number of growing degreeunits to pollen shed and to silk emergence, plant height and stay green.

Table 2C compares Pioneer Brand Hybrid 33M72 and Hybrid 34N43, asimilarly adapted hybrid. The table demonstrates significant differencesbetween Hybrid 33M72 and Hybrid 34N43 which include number of growingdegree units to pollen shed, stay green and resistance to AnthracnoseStalk Rot.

Table 2D compares Pioneer Brand Hybrid 33M72 and Hybrid 33M54, asimilarly adapted hybrid. Significant differences between Hybrid 33M72and Hybrid 33M54 include harvest moisture and number of growing degreeunits to pollen shed and to silk emergence. It is expected that othersignificant differences would be demonstrated if additional traits areobserved.

TABLE 2A HYBRID COMPARISON Variety #1: 33M72 Variety #2: 34H31 YIELDYIELD EGRWTH ESTCNT GDUSHD BU/A 56# BU/A 56# MST PCT SCORE COUNT GDUStat ABS % MN % MN % MN % MN % MN Mean1 191.5 104.1 104.1 96.2 94.3 98.9Mean2 192.6 104.4 99.2 102.6 104.5 99.2 Locs 23 23 25 13 1 8 Reps 23 2325 14 2 9 Diff −1.1 −0.3 −4.9 −6.4 −10.2 −0.4 Prob 0.797 0.883 0.0090.094 0.586 GDUSLK STKCNT PLTHT EARHT STAGRN TSTWT GDU COUNT CM CM SCORELB/BU Stat % MN % MN % MN % MN % MN ABS Mean1 98.6 100.2 101.3 105.9118.2 56.7 Mean2 97.5 101.0 96.8 95.2 84.0 57.8 Locs 6 37 10 10 9 19Reps 7 44 11 11 9 19 Diff 1.1 −0.8 4.5 10.7 34.2 −1.2 Prob 0.151 0.0100.031 0.007 0.083 0.003 GLFSPT SLFBLT ANTROT FUSERS ECBDPE GIBROT SCORESCORE SCORE SCORE % NOT SCORE Stat ABS ABS ABS ABS ABS ABS Mean1 4.8 4.04.5 6.3 97.2 5.0 Mean2 5.5 5.5 4.7 5.8 97.2 7.5 Locs 5 1 3 2 1 1 Reps 72 5 3 1 2 Diff −0.7 −1.5 −0.2 0.5 0.0 −2.5 Prob 0.184 0.742 0.795 BRTSTKERTLPN LRTLPN % NOT % NOT % NOT Stat ABS ABS ABS Mean1 86.1 75.0 100.0Mean2 98.6 95.0 90.0 Locs 2 2 2 Reps 3 2 2 Diff −12.5 −20.0 10.0 Prob0.463 0.626 0.500

TABLE 2B HYBRID COMPARISON Variety #1: 33M72 Variety #2: 33P66 YIELDYIELD EGRWTH ESTCNT GDUSHD BU/A 56# BU/A 56# MST PCT SCORE COUNT GDUStat ABS % MN % MN % MN % MN % MN Mean1 192.3 104.3 104.1 96.2 94.3 98.9Mean2 191.0 103.5 105.9 85.3 104.5 103.9 Locs 22 22 25 13 1 8 Reps 22 2225 14 2 9 Diff 1.3 0.8 1.8 10.8 −10.2 −5.0 Prob 0.795 0.787 0.331 0.0530.001 GDUSLK STKCNT PLTHT EARHT STAGRN TSTWT GDU COUNT CM CM SCORE LB/BUStat % MN % MN % MN % MN % MN ABS Mean1 98.6 100.2 101.3 105.9 118.256.8 Mean2 103.9 100.0 106.3 106.1 88.5 57.0 Locs 6 37 10 10 9 20 Reps 744 11 11 9 20 Diff −5.3 0.3 −5.0 −0.2 29.7 −0.2 Prob 0.002 0.673 0.0000.946 0.024 0.617 GLFSPT SLFBLT ANTROT FUSERS ECBDPE GIBROT SCORE SCORESCORE SCORE % NOT SCORE Stat ABS ABS ABS ABS ABS ABS Mean1 4.8 4.0 4.56.3 97.2 5.0 Mean2 4.4 5.5 3.8 5.0 100.0 4.0 Locs 5 1 3 2 1 1 Reps 7 2 53 1 2 Diff 0.4 −1.5 0.7 1.3 −2.8 1.0 Prob 0.099 0.270 0.500 BRTSTKERTLPN LRTLPN % NOT % NOT % NOT Stat ABS ABS ABS Mean1 86.1 75.0 100.0Mean2 98.6 90.0 95.0 Locs 2 2 2 Reps 3 2 2 Diff −12.5 −15.0 5.0 Prob0.421 0.500 0.500

TABLE 2C HYBRID COMPARISON Variety #1: 33M72 Variety #2: 34N43 YIELDYIELD EGRWTH ESTCNT GDUSHD BU/A 56# BU/A 56# MST PCT SCORE COUNT GDUStat ABS % MN % MN % MN % MN % MN Mean1 190.5 104.0 103.9 96.2 94.3 98.9Mean2 196.2 107.0 103.6 107.8 105.7 96.9 Locs 22 22 24 13 1 8 Reps 22 2224 14 2 9 Diff −5.7 −3.0 −0.3 −11.6 −11.5 2.0 Prob 0.174 0.183 0.8830.036 0.007 GDUSLK STKCNT PLTHT EARHT STAGRN TSTWT GDU COUNT CM CM SCORELB/BU Stat % MN % MN % MN % MN % MN ABS Mean1 98.6 100.2 101.3 105.9118.2 56.6 Mean2 96.8 101.0 95.3 97.5 90.5 56.3 Locs 6 37 10 10 9 19Reps 7 44 11 11 9 19 Diff 1.8 −0.7 5.9 8.4 27.7 0.3 Prob 0.085 0.0530.005 0.009 0.028 0.467 GLFSPT SLFBLT ANTROT FUSERS ECBDPE GIBROT SCORESCORE SCORE SCORE % NOT SCORE Stat ABS ABS ABS ABS ABS ABS Mean1 4.8 4.04.5 6.3 97.2 5.0 Mean2 4.7 6.0 3.3 5.3 98.6 4.0 Locs 5 1 3 2 1 1 Reps 72 5 3 1 2 Diff 0.1 −2.0 1.2 1.0 −1.4 1.0 Prob 0.815 0.020 0.500 BRTSTKERTLPN LRTLPN % NOT % NOT % NOT Stat ABS ABS ABS Mean1 86.1 75.0 100.0Mean2 100.0 95.0 100.0 Locs 2 2 2 Reps 3 2 2 Diff −13.9 −20.0 0.0 Prob0.389 0.626 1.000

TABLE 2D HYBRID COMPARISON Variety #1: 33M72 Variety #2: 33M54 YIELDYIELD EGRWTH ESTCNT GDUSHD BU/A 56# BU/A 56# MST PCT SCORE COUNT GDUStat ABS % MN % MN % MN % MN % MN Mean1 191.5 103.8 104.1 96.2 94.3 98.9Mean2 191.4 103.7 112.7 89.7 103.2 103.2 Locs 20 20 22 13 1 8 Reps 20 2022 14 2 9 Diff 0.0 0.2 8.6 6.5 −8.9 −4.3 Prob 0.997 0.953 0.002 0.1160.012 GDUSLK STKCNT PLTHT EARHT STAGRN TSTWT GDU COUNT CM CM SCORE LB/BUStat % MN % MN % MN % MN % MN ABS Mean1 98.6 100.2 101.3 105.9 118.256.9 Mean2 103.2 100.5 104.3 101.7 117.3 57.5 Locs 6 34 10 10 9 18 Reps7 39 11 11 9 18 Diff −4.6 −0.3 −3.0 4.3 0.8 −0.7 Prob 0.016 0.479 0.0860.314 0.951 0.143 GLFSPT ANTROT ECBDPE BRTSTK ERTLPN LRTLPN SCORE SCORE% NOT % NOT % NOT % NOT Stat ABS ABS ABS ABS ABS ABS Mean1 4.6 5.0 97.286.1 75.0 100.0 Mean2 6.0 6.0 91.7 96.5 70.0 90.0 Locs 4 1 1 2 2 2 Reps5 2 1 3 2 2 Diff −1.4 −1.0 5.6 −10.4 5.0 10.0 Prob 0.011 0.403 0.9100.500Further Embodiments of the Invention

This invention also is directed to methods for producing a maize plantby crossing a first parent maize plant with a second parent maize plantwherein either the first or second parent maize plant is Pioneer Brandhybrid 33M72. In one embodiment the parent hybrid maize plant 33M72 willbe crossed with another maize plant, sibbed, or selfed, to generate aninbred which may be used in the development of additional plants. Inanother embodiment, double haploid methods may be used to generate aninbred plant. Further, this invention is directed to methods forproducing a hybrid 33M72-progeny maize plant by crossing hybrid maizeplant 33M72 with itself or a second maize plant and growing the progenyseed, and repeating the crossing and growing steps with the hybrid maize33M72-progeny plant from 1 to 2 times, 1 to 3 times, 1 to 4 times, or 1to 5 times. Thus, any such methods using the hybrid maize plant 33M72are part of this invention: selfing, sibbing, backcrosses, hybridproduction, crosses to populations, and the like.

All plants produced using hybrid maize plant 33M72 as a parent arewithin the scope of this invention, including plants derived from hybridmaize plant 33M72. Progeny of the breeding methods described herein maybe characterized in any number of ways, such as by traits retained inthe progeny, pedigree and/or molecular markers. Combinations of thesemethods of characterization may be used. This includes varietiesessentially derived from variety 33M72 with the term “essentiallyderived variety” having the meaning ascribed to such term in 7 U.S.C. §2104(a)(3) of the Plant Variety Protection Act, which definition ishereby incorporated by reference. This also includes progeny plant andparts thereof with at least one ancestor that is hybrid maize plant33M72 and more specifically where the pedigree of this progeny includes1, 2, 3, 4, and/or 5 or cross pollinations to a maize plant 33M72, or aplant that has 33M72 as a progenitor. Pedigree is a method used bybreeders of ordinary skill in the art to describe the varieties.Varieties that are more closely related by pedigree are likely to sharecommon genotypes and combinations of phenotypic characteristics. Allbreeders of ordinary skill in the art maintain pedigree records of theirbreeding programs. These pedigree records contain a detailed descriptionof the breeding process, including a listing of all parental lines usedin the breeding process and information on how such line was used. Thus,a breeder of ordinary skill in the art would know if 33M72 were used inthe development of a progeny line, and would also know how many breedingcrosses to a line other than 33M72 were made in the development of anyprogeny line. A progeny line so developed may then be used in crosseswith other, different, maize inbreds to produce first generation (F₁)maize hybrid seeds and plants with superior characteristics.

Specific methods and products produced using hybrid maize plant in plantbreeding are encompassed within the scope of the invention listed above.One such embodiment is the method of crossing hybrid maize plant 33M72with itself to form a homozygous inbred parent line. Hybrid 33M72 wouldbe sib or self pollinated to form a population of progeny plants. Thepopulation of progeny plants produced by this method is also anembodiment of the invention. This first population of progeny plantswill have received all of its alleles from hybrid maize plant 33M72. Theinbreeding process results in homozygous loci being generated and isrepeated until the plant is homozygous at substantially every loci andbecomes an inbred line. Once this is accomplished the inbred line may beused in crosses with other inbred lines, including but not limited toinbred parent lines disclosed herein to generate a first generation ofF1 hybrid plants. One of ordinary skill in the art can utilize breedernotebooks, or molecular methods to identify a particular hybrid plantproduced using an inbred line derived from maize hybrid plant 33M72, inaddition to comparing traits. Any such individual inbred plant is alsoencompassed by this invention.

These embodiments also include use of these methods with transgenic orbackcross conversions of maize hybrid plant 33M72. Another suchembodiment is a method of developing a line genetically similar tohybrid maize plant 33M72 in breeding that involves the repeatedbackcrossing of an inbred parent of, or an inbred line derived from,hybrid maize plant 33M72 to another different maize plant any number oftimes. Using backcrossing methods, or even the tissue culture andtransgenic methods described herein, the backcross conversion methodsdescribed herein, or other breeding methods known to one of ordinaryskill in the art, one can develop individual plants, plant cells, andpopulations of plants that retain at least 25%, 30%, 35%, 40%, 45%, 50%,55%, 60%, 65%, 70%, 75%, 79%, 80%, 82%, 83%, 84%, 85%, 86%, 87%, 88%,89%, 90%, 91%, 92%, 93%, 94%, 96%, 97%, 98%, 99% or 99.5% geneticsimilarity or identity to maize hybrid plant 33M72. The percentage ofthe genetics retained in the progeny may be measured by either pedigreeanalysis or through the use of genetic techniques such as molecularmarkers or electrophoresis. In pedigree analysis, on average 50% of thestarting germplasm would be passed to the progeny line after one crossto a different line, 25% after another cross to a different line, and soon. Molecular markers could also be used to confirm and/or determine thepedigree of the progeny line. The inbred parent would then be crossed toa second inbred parent of or derived from hybrid maize plant 33M72 tocreate hybrid maize plant 33M72 with additional beneficial traits suchas transgenes or backcross conversions.

One method for producing a line derived from hybrid maize plant is asfollows. One of ordinary skill in the art would obtain hybrid maizeplant 33M72 and cross it with another variety of maize, such as an eliteinbred variety. The F1 seed derived from this cross would be grown toform a population. The nuclear genome of the F1 would be made-up of 50%of hybrid maize plant 33M72 and 50% of the other elite variety. The F1seed would be grown and allowed to self, thereby forming F2 seed. Onaverage the F2 seed nuclear genome would have derived 50% of its allelesfrom the parent hybrid plant 33M72 and 50% from the other maize variety,but various individual plants from the population would have a muchgreater percentage of their alleles derived from the parent maize hybridplant (Wang J. and R. Bernardo, 2000, Crop Sci. 40:659–665 and Bernardo,R. and A. L. Kahler, 2001, Theor. Appl. Genet 102: 986–992). Molecularmarkers of 33M72, or its parents identified from routine screening ofthe deposited samples herein could be used to select and retain thoselines with high similarity to 33M72. The F2 seed would be grown andselection of plants would be made based on visual observation, markersand/or measurement of traits. The traits used for selection may be any33M72 trait described in this specification, including the hybrid maizeplant 33M72 traits of yield, overall agronomics, stalk strength, staygreen, late season root strength, Corn Rootworm resistance, Goss' Wiltresistance, resistance to Northern Leaf Blight, Diplodia Ear Rotresistance, resistance to Fusarium Ear and Kernel Rot, and isparticularly suited to the Central Corn Belt, Eastern, and Western areasof the United States.

Such traits may also be the good general or specific combining abilityof 33M72, including its ability to produce backcross conversions, orother hybrids. The 33M72 progeny plants that exhibit one or more of thedesired 33M72 traits, such as those listed above, would be selected andeach plant would be harvested separately. This F3 seed from each plantwould be grown in individual rows and allowed to self. Then selectedrows or plants from the rows would be harvested individually. Theselections would again be based on visual observation, markers and/ormeasurements for desirable traits of the plants, such as one or more ofthe desirable 33M72 traits listed above.

The process of growing and selection would be repeated any number oftimes until a 33M72 progeny plant is obtained. The 33M72 progeny inbredplant would contain desirable traits in hybrid combination derived fromhybrid plant 33M72. The resulting progeny line would benefit from theefforts of the inventor(s), and would not have existed but for theinventor(s) work in creating 33M72. Another embodiment of the inventionis a 33M72 progeny plant that has received the desirable 33M72 traitslisted above through the use of 33M72, which traits were not exhibitedby other plants used in the breeding process.

The previous example can be modified in numerous ways, for instanceselection may or may not occur at every selfing generation, the hybridmay immediately be selfed without crossing to another plant, selectionmay occur before or after the actual self-pollination process occurs, orindividual selections may be made by harvesting individual ears, plants,rows or plots at any point during the breeding process described. Inaddition, double haploid breeding methods may be used at any step in theprocess. The population of plants produced at each and any cycle ofbreeding is also an embodiment of the invention, and on average eachsuch population would predictably consist of plants containingapproximately 50% of its genes from inbred parents of maize hybrid 33M72in the first breeding cycle, 25% of its genes from inbred parents ofmaize hybrid 33M72 in the second breeding cycle, 12.5% of its genes frominbred parents of maize hybrid 33M72 in the third breeding cycle, 6.25%in the fourth breeding cycle, 3.125% in the fifth breeding cycle, and soon. In each case the use of 33M72 provides a substantial benefit. Thelinkage groups of 33M72 would be retained in the progeny lines, andsince current estimates of the maize genome size is about 50,000–80,000genes (Xiaowu, Gai et al., Nucleic Acids Research, 2000, Vol. 28, No. 1,94–96), in addition to a large amount of non-coding DNA that impactsgene expression, it provides a significant advantage to use 33M72 asstarting material to produce a line that retains desired genetics ortraits of 33M72.

Another embodiment of this invention is the method of obtaining asubstantially homozygous 33M72 progeny plant by obtaining a seed fromthe cross of 33M72 and another maize plant and applying double haploidmethods to the F1 seed or F1 plant or to any successive filialgeneration. Such methods substantially decrease the number ofgenerations required to produce an inbred with similar genetics orcharacteristics to 33M72.

A further embodiment of the invention is a backcross conversion of 33M72obtained by crossing inbred parent plants of hybrid maize plant 33M72,which comprise the backcross conversion. For a dominant or additivetrait at least one of the inbred parents would include backcrossconversion in its genome. For a recessive trait, each parent wouldinclude the backcross conversion in its genome. In each case theresultant hybrid maize plant 33M72 obtained from the cross of theparents includes a backcross conversion or transgene.

A backcross conversion of 33M72 occurs when DNA sequences are introducedthrough traditional (non-transformation) breeding techniques, such asbackcrossing (Hallauer et al., 1988), with a parent of 33M72 utilized asthe recurrent parent. Both naturally occurring and transgenic DNAsequences may be introduced through backcrossing techniques. The termbackcross conversion is also referred to in the art as a single locusconversion. A backcross conversion may produce a plant with a trait orlocus conversion in at least one or more backcrosses, including at least2 crosses, at least 3 crosses, at least 4 crosses, at least 5 crossesand the like. Molecular marker assisted breeding or selection may beutilized to reduce the number of backcrosses necessary to achieve thebackcross conversion. For example, see Openshaw, S. J. et al.,Marker-assisted Selection in Backcross Breeding. In: ProceedingsSymposium of the Analysis of Molecular Data, August 1994, Crop ScienceSociety of America, Corvallis, Oreg., where it is demonstrated that abackcross conversion can be made in as few as two backcrosses.

The complexity of the backcross conversion method depends on the type oftrait being transferred (single genes or closely linked genes as vs.unlinked genes), the level of expression of the trait, the type ofinheritance (cytoplasmic or nuclear) and the types of parents includedin the cross. It is understood by those of ordinary skill in the artthat for single gene traits that are relatively easy to classify, thebackcross method is effective and relatively easy to manage. (SeeHallauer et al. in Corn and Corn Improvement, Sprague and Dudley, ThirdEd. 1998). Desired traits that may be transferred through backcrossconversion include, but are not limited to, waxy starch, sterility(nuclear and cytoplasmic), fertility restoration, grain color (white),nutritional enhancements, drought tolerance, nitrogen utilization,altered fatty acid profile, increased digestibility, low phytate,industrial enhancements, disease resistance (bacterial, fungal orviral), insect resistance, herbicide resistance and yield enhancements.In addition, an introgression site itself, such as an FRT site, Lox siteor other site specific integration site, may be inserted by backcrossingand utilized for direct insertion of one or more genes of interest intoa specific plant variety. The trait of interest is transferred from thedonor parent to the recurrent parent, in this case, an inbred parent ofthe maize plant disclosed herein. In some embodiments of the invention,the number of loci that may be backcrossed into 33M72 is at least 1, 2,3, 4, or 5 and/or no more than 6, 5, 4, 3, or 2. A single loci maycontain several transgenes, such as a transgene for disease resistancethat, in the same expression vector, also contains a transgene forherbicide resistance. The gene for herbicide resistance may be used as aselectable marker and/or as a phenotypic trait. A single locusconversion of site specific integration system allows for theintegration of multiple genes at the converted loci.

The backcross conversion may result from either the transfer of adominant allele or a recessive allele. Selection of progeny containingthe trait of interest is accomplished by direct selection for a traitassociated with a dominant allele. Transgenes transferred viabackcrossing typically function as a dominant single gene trait and arerelatively easy to classify. Selection of progeny for a trait that istransferred via a recessive allele, such as the waxy starchcharacteristic, requires growing and selfing the first backcrossgeneration to determine which plants carry the recessive alleles.Recessive traits may require additional progeny testing in successivebackcross generations to determine the presence of the locus ofinterest. The last backcross generation is usually selfed to give purebreeding progeny for the gene(s) being transferred, although a backcrossconversion with a stably introgressed trait may also be maintained byfurther backcrossing to the recurrent parent with selection for theconverted trait.

Along with selection for the trait of interest, progeny are selected forthe phenotype of the recurrent parent. While occasionally additionalpolynucleotide sequences or genes may be transferred along with thebackcross conversion, the backcross conversion line “fits into the samehybrid combination as the recurrent parent inbred line and contributesthe effect of the additional gene added through the backcross.” Poehlmanet al. (1995, pg. 334). A progeny comprising at least 95%, 96%, 97%,98%, 99%, 99.5% and 99.9% genetic identity to hybrid 33M72 andcomprising the backcross conversion trait or traits of interest, isconsidered to be a backcross conversion of hybrid 33M72. It has beenproposed that in general there should be at least four backcrosses whenit is important that the recovered lines be essentially identical to therecurrent parent except for the characteristic being transferred (Fehr1987, Principles of Cultivar Development). However, as noted above, thenumber of backcrosses necessary can be reduced with the use of molecularmarkers. Other factors, such as a genetically similar donor parent, mayalso reduce the number of backcrosses necessary.

Hybrid seed production requires elimination or inactivation of pollenproduced by the female inbred parent. Incomplete removal or inactivationof the pollen provides the potential for self-pollination. A reliablemethod of controlling male fertility in plants offers the opportunityfor improved seed production. It should be understood that the plantcan, through routine manipulation by detasseling, cytoplasmic genes,nuclear genes, or other factors, be produced in a male-sterile form. Theterm manipulated to be male sterile refers to the use of any availabletechniques to produce a male sterile version of maize line 33M72. Themale sterility may be either partial or complete male sterility.

Hybrid maize seed is often produced by a male sterility systemincorporating manual or mechanical detasseling. Alternate strips of twomaize inbreds are planted in a field, and the pollen-bearing tassels areremoved from one of the inbreds (female). Provided that there issufficient isolation from sources of foreign maize pollen, the ears ofthe detasseled inbred will be fertilized only from the other inbred(male), and the resulting seed is therefore hybrid and will form hybridplants.

Such embodiments are also within the scope of the present claims. Thisinvention includes hybrid maize seed of 33M72 and the hybrid maize plantproduced therefrom. The foregoing was set forth by way of example and isnot intended to limit the scope of the invention.

This invention is also directed to the use of hybrid maize plant 33M72in tissue culture. As used herein, the term plant includes plantprotoplasts, plant cell tissue cultures from which maize plants can beregenerated, plant calli, plant clumps, and plant cells that are intactin plants, or parts of plants, such as embryos, pollen, ovules, flowers,kernels, ears, cobs, leaves, seeds, husks, stalks, roots, root tips,anthers, silk and the like. As used herein the phrase “growing the seed”or “grown from the seed” includes embryo rescue, isolation of cells fromseed for use in tissue culture, as well as traditional growing methods.

Duncan, Williams, Zehr, and Widholm, Planta, (1985) 165:322–332 reflectsthat 97% of the plants cultured which produced callus were capable ofplant regeneration. Subsequent experiments with both inbreds and hybridsproduced 91% regenerable callus which produced plants. In a furtherstudy in 1988, Songstad, Duncan & Widholm in Plant Cell Reports (1988),7:262–265 reports several media additions which enhance regenerabilityof callus of two inbred lines. Other published reports also indicatedthat “nontraditional” tissues are capable of producing somaticembryogenesis and plant regeneration. K. P. Rao, et al., Maize GeneticsCooperation Newsletter, 60:64–65 (1986), refers to somatic embryogenesisfrom glume callus cultures and B. V. Conger, et al., Plant Cell Reports,6:345–347 (1987) indicates somatic embryogenesis from the tissuecultures of maize leaf segments. Thus, it is clear from the literaturethat the state of the art is such that these methods of obtaining plantsare, and were, “conventional” in the sense that they are routinely usedand have a very high rate of success.

Tissue culture of maize, including tassel/anther culture, is describedin U.S. Application 2002/0062506A1 and European Patent Application,Publication No. 160,390, each of which are incorporated herein byreference. Maize tissue culture procedures are also described in Greenand Rhodes, “Plant Regeneration in Tissue Culture of Maize,” Maize forBiological Research (Plant Molecular Biology Association,Charlottesville, Virginia 1982, at 367–372) and in Duncan, et al., “TheProduction of Callus Capable of Plant Regeneration from Immature Embryosof Numerous Zea Mays Genotypes,” 165 Planta 322–332 (1985). Thus,another aspect of this invention is to provide cells which upon growthand differentiation produce maize plants having the genotype and/orphysiological and morphological characteristics of hybrid maize plant33M72.

The utility of hybrid maize plant 33M72 also extends to crosses withother species. Commonly, suitable species will be of the familyGraminaceae, and especially of the genera Zea, Tripsacum, Coix,Schlerachne, Polytoca, Chionachne, and Trilobachne, of the tribeMaydeae. Potentially suitable for crosses with 33M72 may be the variousvarieties of grain sorghum, Sorghum bicolor (L.) Moench.

Transformation of Maize

The advent of new molecular biological techniques has allowed theisolation and characterization of genetic elements with specificfunctions, such as encoding specific protein products. Scientists in thefield of plant biology developed a strong interest in engineering thegenome of plants to contain and express foreign genetic elements, oradditional, or modified versions of native or endogenous geneticelements in order to alter the traits of a plant in a specific manner.Any DNA sequences, whether from a different species or from the samespecies, that are inserted into the genome using transformation arereferred to herein collectively as “transgenes”. In some embodiments ofthe invention, a transformed variant of 33M72 may contain at least onetransgene but could contain at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10and/or no more than 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2.Over the last fifteen to twenty years several methods for producingtransgenic plants have been developed, and the present invention alsorelates to transformed versions of the claimed hybrid 33M72 as well ascombinations thereof.

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols. See, forexample, Miki et al., “Procedures for Introducing Foreign DNA intoPlants” in Methods in Plant Molecular Biology and Biotechnology, Glick,B. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton, 1993) pp.67–88 and Armstrong, “The First Decade of Maize Transformation: A Reviewand Future Perspective” (Maydica 44:101–109, 1999). In addition,expression vectors and in vitro culture methods for plant cell or tissuetransformation and regeneration of plants are available. See, forexample, Gruber et al., “Vectors for Plant Transformation” in Methods inPlant Molecular Biology and Biotechnology, Glick, B. R. and Thompson, J.E. Eds. (CRC Press, Inc., Boca Raton, 1993) pp. 89–119. See U.S. Pat.No. 6,118,055, which is herein incorporated by reference.

The most prevalent types of plant transformation involve theconstruction of an expression vector. Such a vector comprises a DNAsequence that contains a gene under the control of or operatively linkedto a regulatory element, for example a promoter. The vector may containone or more genes and one or more regulatory elements.

A genetic trait which has been engineered into the genome of aparticular maize plant using transformation techniques, could be movedinto the genome of another line using traditional breeding techniquesthat are well known in the plant breeding arts. These lines can then becrossed to generate a hybrid maize plant such as hybrid maize plant33M72 which comprises a transgene. For example, a backcrossing approachis commonly used to move a transgene from a transformed maize plant toan elite inbred line, and the resulting progeny would then comprise thetransgene(s). Also, if an inbred line was used for the transformationthen the transgenic plants could be crossed to a different inbred inorder to produce a transgenic hybrid maize plant. As used herein,“crossing” can refer to a simple X by Y cross, or the process ofbackcrossing, depending on the context.

Various genetic elements can be introduced into the plant genome usingtransformation. These elements include but are not limited to genes;coding sequences; inducible, constitutive, and tissue specificpromoters; enhancing sequences; and signal and targeting sequences. Forexample, see the traits, genes and transformation methods listed in U.S.Pat. No. 6,284,953, which is herein incorporated by reference.

With transgenic plants according to the present invention, a foreignprotein can be produced in commercial quantities. Thus, techniques forthe selection and propagation of transformed plants, which are wellunderstood in the art, yield a plurality of transgenic plants that areharvested in a conventional manner, and a foreign protein then can beextracted from a tissue of interest or from total biomass. Proteinextraction from plant biomass can be accomplished by known methods whichare discussed, for example, by Heney and Orr, Anal. Biochem. 114: 92–6(1981). In one embodiment, the biomass of interest is seed.

A genetic map can be generated, primarily via conventional RestrictionFragment Length Polymorphisms (RFLP), Polymerase Chain Reaction (PCR)analysis, Simple Sequence Repeats (SSR) and Single NucleotidePolymorphisms (SNP) which identifies the approximate chromosomallocation of the integrated DNA molecule. For exemplary methodologies inthis regard, see Glick and Thompson, METHODS IN PLANT MOLECULAR BIOLOGYAND BIOTECHNOLOGY 269–284 (CRC Press, Boca Raton, 1993).

Wang et al. discuss “Large Scale Identification, Mapping and Genotypingof Single-Nucleotide Polymorphisms in the Human Genome”, Science, 280:1077–1082, 1998, and similar capabilities are becoming increasinglyavailable for the corn genome. Map information concerning chromosomallocation is useful for proprietary protection of a subject transgenicplant. If unauthorized propagation is undertaken and crosses made withother germplasm, the map of the integration region can be compared tosimilar maps for suspect plants to determine if the latter have a commonparentage with the subject plant. Map comparisons would involvehybridizations, RFLP, PCR, SSR and sequencing, all of which areconventional techniques. SNPs may also be used alone or in combinationwith other techniques.

Likewise, by means of the present invention, plants can be geneticallyengineered to express various phenotypes of agronomic interest. Throughthe transformation of maize the expression of genes can be modulated toenhance disease resistance, insect resistance, herbicide resistance,agronomic traits, grain quality and other traits. Transformation canalso be used to insert DNA sequences which control or help controlmale-sterility. DNA sequences native to maize as well as non-native DNAsequences can be transformed into maize and used to modulate levels ofnative or non-native proteins. Various promoters, targeting sequences,enhancing sequences, and other DNA sequences can be inserted into themaize genome for the purpose of modulating the expression of proteins.Reduction of the activity of specific genes (also known as genesilencing, or gene suppression) is desirable for several aspects ofgenetic engineering in plants.

Many techniques for gene silencing are well known to one of skill in theart, including but not limited to antisense technology (see, e.g.,Sheehy et al. (1988) PNAS USA 85: 8805–8809; and U.S. Pat. Nos.5,107,065; 5,453, 566; and 5,759,829); co-suppression (e.g., Taylor(1997) Plant Cell 9:1245; Jorgensen (1990) Trends Biotech.8(12):340–344; Flavell (1994) PNAS USA 91:3490–3496; Finnegan et al.(1994) Bio/Technology 12: 883–888; and Neuhuber et al. (1994) Mol. Gen.Genet. 244:230–241); RNA interference (Napoli et al. (1990) Plant Cell2:279–289; U.S. Pat. No. 5,034,323; Sharp (1999) Genes Dev. 13:139–141;Zamore et al. (2000) Cell 101:25–33; and Montgomery et al. (1998) PNASUSA 95:15502–15507), virus-induced gene silencing (Burton, et al. (2000)Plant Cell 12:691–705; and Baulcombe (1999) Curr. Op. Plant Bio.2:109–113); target-RNA-specific ribozymes (Haseloff et al. (1988) Nature334: 585–591); hairpin structures (Smith et al. (2000) Nature407:319–320; WO 99/53050; and WO 98/53083); ribozymes (Steinecke et al.(1992) EMBO J. 11:1525; and Perriman et al. (1993) Antisense Res. Dev.3: 253); oligonucleotide mediated targeted modification (e.g., WO03/076574 and WO 99/25853); Zn-finger targeted molecules (e.g., WO01/52620; WO 03/048345; and WO 00/42219); and other methods orcombinations of the above methods known to those of skill in the art.

Exemplary transgenes useful for genetic engineering include, but are notlimited to, those categorized below.

1. Transgenes that Confer Resistance to Pests or Disease and thatEncode:

(A) Plant disease resistance genes. Plant defenses are often activatedby specific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant variety can be transformed with clonedresistance gene to engineer plants that are resistant to specificpathogen strains. See, for example Jones et al., Science 266: 789 (1994)(cloning of the tomato Cf-9 gene for resistance to Cladosporium fulvum);Martin et al., Science 262: 1432 (1993) (tomato Pto gene for resistanceto Pseudomonas syringae pv. tomato encodes a protein kinase); Mindrinoset al., Cell 78: 1089 (1994) (Arabidopsis RSP2 gene for resistance toPseudomonas syringae). A plant resistant to a disease is one that ismore resistant to a pathogen as compared to the wild type plant.

(B) A Bacillus thuringiensis protein, a derivative thereof or asynthetic polypeptide modeled thereon. See, for example, Geiser et al.,Gene 48: 109 (1986), who disclose the cloning and nucleotide sequence ofa Bt delta-endotoxin gene. Moreover, DNA molecules encodingdelta-endotoxin genes can be purchased from American Type CultureCollection (Rockville, Md.), for example, under ATCC Accession Nos.40098, 67136, 31995 and 31998. Other examples of Bacillus thuringiensistransgenes being genetically engineered are given in the followingpatents and hereby are incorporated by reference for this purpose: U.S.Pat. Nos. 5,188,960; 5,689,052; 5,880,275; WO 91/114778; WO 99/31248; WO01/12731; WO 99/24581; WO 97/40162 and U.S. patent Ser. Nos. 10/032,717;10/414,637; and 10/606,320.

(C) An insect-specific hormone or pheromone such as an ecdysteroid andjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof. See, for example, the disclosure byHammock et al., Nature 344: 458 (1990), of baculovirus expression ofcloned juvenile hormone esterase, an inactivator of juvenile hormone.

(D) An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Regan, J. Biol. Chem. 269: 9 (1994) (expression cloningyields DNA coding for insect diuretic hormone receptor), and Pratt etal., Biochem. Biophys. Res. Comm. 163: 1243 (1989) (an allostatin isidentified in Diploptera puntata). See also U.S. Pat. No. 5,266,317 toTomalski et al., who disclose genes encoding insect-specific, paralyticneurotoxins.

(E) An enzyme responsible for an hyperaccumulation of a monterpene, asesquiterpene, a steroid, hydroxamic acid, a phenylpropanoid derivativeor another non-protein molecule with insecticidal activity.

(F) An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase and a glucanase, whether natural or synthetic. See PCTApplication WO 93/02197 in the name of Scott et al., which discloses thenucleotide sequence of a callase gene. DNA molecules which containchitinase-encoding sequences can be obtained, for example, from the ATCCunder Accession Nos. 39637 and 67152. See also Kramer et al., InsectBiochem. Molec. Biol. 23: 691 (1993), who teach the nucleotide sequenceof a cDNA encoding tobacco hookworm chitinase, and Kawalleck et al.,Plant Molec. Biol. 21: 673 (1993), who provide the nucleotide sequenceof the parsley ubi4-2 polyubiquitin gene.

(G) A molecule that stimulates signal transduction. For example, see thedisclosure by Botella et al., Plant Molec. Biol. 24: 757 (1994), ofnucleotide sequences for mung bean calmodulin cDNA clones, and Griess etal., Plant Physiol. 104: 1467 (1994), who provide the nucleotidesequence of a maize calmodulin cDNA clone.

(H) A hydrophobic moment peptide. See PCT Application WO 95/16776(disclosure of peptide derivatives of Tachyplesin which inhibit fungalplant pathogens) and PCT Application WO 95/18855 (teaches syntheticantimicrobial peptides that confer disease resistance), the respectivecontents of which are hereby incorporated by reference for this purpose.

(I) A membrane permease, a channel former or a channel blocker. Forexample, see the disclosure by Jaynes et al., Plant Sci. 89: 43 (1993),of heterologous expression of a cecropin-beta lytic peptide analog torender transgenic tobacco plants resistant to Pseudomonas solanacearum.

(J) A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. See Beachy et al., Ann. Rev. Phytopathol.28: 451 (1990). Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,tobacco streak virus, potato virus X, potato virus Y, tobacco etchvirus, tobacco rattle virus and tobacco mosaic virus. Id.

(K) An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect. Cf.Taylor et al., Abstract #497, SEVENTH INT'L SYMPOSIUM ON MOLECULARPLANT-MICROBE INTERACTIONS (Edinburgh, Scotland, 1994) (enzymaticinactivation in transgenic tobacco via production of single-chainantibody fragments).

(L) A virus-specific antibody. See, for example, Taviadoraki et al.,Nature 366: 469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

(M) A developmental-arrestive protein produced in nature by a pathogenor a parasite. Thus, fungal endo alpha-1,4-D-polygalacturonasesfacilitate fungal colonization and plant nutrient release bysolubilizing plant cell wall homo-alpha-1,4-D-galacturonase. See Lamb etal., Bio/Technology 10: 1436 (1992). The cloning and characterization ofa gene which encodes a bean endopolygalacturonase-inhibiting protein isdescribed by Toubart et al., Plant J. 2: 367 (1992).

(N) A developmental-arrestive protein produced in nature by a plant. Forexample, Logemann et al., Bio/Technology 10: 305 (1992), have shown thattransgenic plants expressing the barley ribosome-inactivating gene havean increased resistance to fungal disease.

(O) Genes involved in the Systemic Acquired Resistance (SAR) Responseand/or the pathogenesis related genes. Briggs, S., Current Biology, 5(2)(1995).

(P) Antifungal genes (Cornelissen and Melchers, Pl. Physiol. 101:709–712, (1993) and Parijs et al., Planta 183:258–264, (1991) andBushnell et al., Can. J. of Plant Path. 20(2):137–149 (1998).

(Q) Detoxification genes, such as for fumonisin, beauvericin,moniliformin and zearalenone and their structurally related derivatives.For example, see U.S. Pat. No. 5,792,931.

(R) Cystatin and cysteine proteinase inhibitors.

(S) Defensin genes. See WO 03/000863.

(T) Genes conferring resistance to nematodes. See WO 03/033651 and Urwinet al., Planta 204:472–479 (1998).

2. Transgenes that Confer Resistance to a Herbicide, for Example:

(A) A herbicide that inhibits the growing point or meristem, such as animidazolinone or a sulfonylurea. Exemplary genes in this category codefor mutant ALS and AHAS enzyme as described, for example, by Lee et al.,EMBO J. 7: 1241 (1988), and Miki et al., Theor. Appl. Genet. 80: 449(1990), respectively. See also, U.S. Pat. Nos. 5,605,011; 5,013,659;5,141,870; 5,767,361; 5,731,180; 5,304,732; 4,761,373; 5,331,107;5,928,937; and 5,378,824; and international publication WO 96/33270,which are incorporated herein by reference in their entireties for thispurpose.

(B) Glyphosate (resistance imparted by mutant5-enolpyruvl-3-phosphikimate synthase (EPSP), and aroA genes,respectively) and other phosphono compounds such as glufosinate(phosphinothricin acetyl transferase (PAT) and Streptomyceshygroscopicus phosphinothricin acetyl transferase (bar) genes), andpyridinoxy or phenoxy proprionic acids and cycloshexones (ACCaseinhibitor-encoding genes). See, for example, U.S. Pat. No. 4,940,835 toShah et al., which discloses the nucleotide sequence of a form of EPSPSwhich can confer glyphosate resistance. U.S. Pat. No. 5,627,061 to Barryet al. also describes genes encoding EPSPS enzymes. See also U.S. Pat.Nos. 6,248,876 B1; 6,040,497; 5,804,425; 5,633,435; 5,145,783;4,971,908; 5,312,910; 5,188,642; 4,940,835; 5,866,775; 6,225,114 B1;6,130,366; 5,310,667; 4,535,060; 4,769,061; 5,633,448; 5,510,471; Re.36,449; RE 37,287 E; and 5,491,288; and international publications WO97/04103; WO 97/04114; WO 00/66746; WO 01/66704; WO 00/66747 and WO00/66748, which are incorporated herein by reference in their entiretiesfor this purpose. Glyphosate resistance is also imparted to plants thatexpress a gene that encodes a glyphosate oxido-reductase enzyme asdescribed more fully in U.S. Pat. Nos. 5,776,760 and 5,463,175, whichare incorporated herein by reference in their entireties for thispurpose. In addition glyphosate resistance can be imparted to plants bythe over expression of genes encoding glyphosate N-acetyltransferase.See, for example, U.S. Application Ser. Nos. 60/244,385; 60/377,175 and60/377,719.

A DNA molecule encoding a mutant aroA gene can be obtained under ATCCAccession No. 39256, and the nucleotide sequence of the mutant gene isdisclosed in U.S. Pat. No. 4,769,061 to Comai. European PatentApplication No. 0 333 033 to Kumada et al. and U.S. Pat. No. 4,975,374to Goodman et al. disclose nucleotide sequences of glutamine synthetasegenes which confer resistance to herbicides such as L-phosphinothricin.The nucleotide sequence of a phosphinothricin-acetyl-transferase gene isprovided in European Patent No. 0 242 246 and 0 242 236 to Leemans etal. De Greef et al., Bio/Technology 7: 61 (1989), describe theproduction of transgenic plants that express chimeric bar genes codingfor phosphinothricin acetyl transferase activity. See also, U.S. Pat.Nos. 5,969,213; 5,489,520; 5,550,318; 5,874,265; 5,919,675; 5,561,236;5,648,477; 5,646,024; 6,177,616 B1; and 5,879,903, which areincorporated herein by reference in their entireties for this purpose.Exemplary genes conferring resistance to phenoxy proprionic acids andcycloshexones, such as sethoxydim and haloxyfop, are the Acc1-S1,Acc1-S2 and Acc1-S3 genes described by Marshall et al., Theor. Appl.Genet. 83: 435 (1992).

(C) A herbicide that inhibits photosynthesis, such as a triazine (psbAand gs+ genes) and a benzonitrile (nitrilase gene). Przibilla et al.,Plant Cell 3: 169 (1991), describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker, andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441 and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes et al., Biochem. J. 285:173 (1992).

(D) Acetohydroxy acid synthase, which has been found to make plants thatexpress this enzyme resistant to multiple types of herbicides, has beenintroduced into a variety of plants (see, e.g., Hattori et al. (1995)Mol Gen Genet 246:419). Other genes that confer tolerance to herbicidesinclude: a gene encoding a chimeric protein of rat cytochrome P4507A1and yeast NADPH-cytochrome P450 oxidoreductase (Shiota et al. (1994)Plant Physiol. 106(1):17–23), genes for glutathione reductase andsuperoxide dismutase (Aono et al. (1995) Plant Cell Physiol. 36:1687,and genes for various phosphotransferases (Datta et al. (1992) PlantMol. Biol. 20: 619).

(E) Protoporphyrinogen oxidase (protox) is necessary for the productionof chlorophyll, which is necessary for all plant survival. The protoxenzyme serves as the target for a variety of herbicidal compounds. Theseherbicides also inhibit growth of all the different species of plantspresent, causing their total destruction. The development of plantscontaining altered protox activity which are resistant to theseherbicides are described in U.S. Pat. Nos. 6,288,306 B1; 6,282,837 B1;and 5,767,373; and international publication WO 01/12825, which areincorporated herein by reference in their entireties.

3. Transgenes that Confer or Contribute to a Grain Trait, such as:

(A) Modified fatty acid metabolism, for example, by

-   -   (1) Transforming a plant with an antisense gene of stearoyl-ACP        desaturase to increase stearic acid content of the plant. See        Knultzon et al., Proc. Natl. Acad. Sci. USA 89: 2624 (1992),    -   (2) Elevating oleic acid via FAD-2 gene modification and/or        decreasing linolenic acid via FAD-3 gene modification (see U.S.        Pat. Nos. 6,063,947; 6,323,392; and WO 93/11245),    -   (3) Altering conjugated linolenic or linoleic acid content, such        as in WO 01/12800,    -   (4) Modifying LEC1, AGP, Dek1, Superal1, thioredoxin, and/or a        gamma zein knock out or mutant such as cs27 or TUSC 27. For        example, see WO 02/42424, WO 98/22604, WO 03/011015, U.S. Pat.        No. 6,423,886 and Rivera-Madrid, R. et. al., Proc. Natl. Acad.        Sci. 92:5620–5624 (1995).

(B) Decreased phytate content, for example, by the

-   -   (1) Introduction of a phytase-encoding gene would enhance        breakdown of phytate, adding more free phosphate to the        transformed plant. For example, see Van Hartingsveldt et al.,        Gene 127: 87 (1993), for a disclosure of the nucleotide sequence        of an Aspergillus niger phytase gene.    -   (2) Introduction of a gene that reduces phytate content. In        maize, this, for example, could be accomplished, by cloning and        then re-introducing DNA associated with one or more of the        alleles, such as the LPA alleles, identified in maize mutants        characterized by low levels of phytic acid, such as in Raboy et        al., Maydica 35: 383 (1990) and/or by altering inositol kinase        activity as in WO 02/059324, U.S. Application No. 2003/0009011,        WO 03/027243, U.S. Application No. 2003/0079247 and WO 99/05298.

(C) Modified carbohydrate composition effected, for example, bytransforming plants with a gene coding for an enzyme that alters thebranching pattern of starch. See Shiroza et al., J. Bacteriol. 170: 810(1988) (nucleotide sequence of Streptococcus mutans fructosyltransferasegene), Steinmetz et al., Mol. Gen. Genet. 200: 220 (1985) (nucleotidesequence of Bacillus subtilis levansucrase gene), Pen et al.,Bio/Technology 10: 292 (1992) (production of transgenic plants thatexpress Bacillus licheniformis alpha-amylase), Elliot et al., PlantMolec. Biol. 21: 515 (1993) (nucleotide sequences of tomato invertasegenes), Søgaard et al., J. Biol. Chem. 268: 22480 (1993) (site-directedmutagenesis of barley alpha-amylase gene), and Fisher et al., PlantPhysiol. 102: 1045 (1993) (maize endosperm starch branching enzyme II).The fatty acid modification genes mentioned above may also be used toeffect starch content and/or composition through the interrelationshipof the starch and oil pathways.

(D) Altered antioxidant content or composition, such as alteration oftocopherol or tocotrienols. For example, see WO 00/68393 involving themanipulation of antioxidant levels through alteration of a phytl prenyltransferase and WO 03/082899 through alteration of a homogentisategeranyl geranyl transferase.

(E) Improved digestibility and/or starch extraction through modificationof UDP-D-xylose 4-epimerase, Fragile 1 and 2, Ref1, HCHL, C4H, such asin WO 99/10498.

4. Genes that Control Male-Sterility:

(A) Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN-Ac-PPT (WO 01/29237).

(B) Introduction of various stamen-specific promoters (WO 92/13956, WO92/13957).

(C) Introduction of the barnase and the barstar gene (Paul et al., PlantMol. Biol. 19:611–622, 1992).

5. Genes that create a site for site specific DNA integration. Thisincludes the introduction of FRT sites that may be used in the FLP/FRTsystem and/or Lox sites that may be used in the Cre/Loxp system. Forexample, see Lyznik, et al., Site-Specific Recombination for GeneticEngineering in Plants, Plant Cell Rep (2003) 21:925–932 which is herebyincorporated by reference. Other systems that may be used include theGin recombinase of phage Mu (Maeser et al., 1991), the Pin recombinaseof E. coli (Enomoto et al., 1983), and the R/RS system of the pSR1plasmid (Araki et al., 1992).6. Genes that affect growth characteristics, such as drought toleranceand nitrogen utilization. For example, see WO 00/73475 where water useefficiency is modulated through alteration of malate.Genetic Marker Profile Through SSR

The present invention comprises a hybrid corn plant which ischaracterized by the molecular and physiological data presented hereinand in the representative sample of said hybrid and of the inbredparents of said hybrid deposited with the ATCC.

To select and develop a superior hybrid, it is necessary to identify andselect genetically unique individuals that occur in a segregatingpopulation. The segregating population is the result of a combination ofcrossover events plus the independent assortment of specificcombinations of alleles at many gene loci that results in specific andunique genotypes. Once such a line is developed its value to society issubstantial since it is important to advance the germplasm base as awhole in order to maintain or improve traits such as yield, diseaseresistance, pest resistance and plant performance in extreme weatherconditions. Backcross trait conversions are routinely used to add ormodify one or a few traits of such a line and this further enhances itsvalue and usefulness to society. The genetic variation among individualprogeny of a breeding cross allows for the identification of rare andvaluable new genotypes. Once identified, it is possible to utilizeroutine and predictable breeding methods to develop progeny that retainthe rare and valuable new genotypes developed by the initial breeder.

Phenotypic traits exhibited by 33M72 can be used to characterize thegenetic contribution of 33M72 to progeny lines developed through the useof 33M72. Quantitative traits including, but not limited to, yield,maturity, stay green, root lodging, stalk lodging, and early growth aretypically governed by multiple genes at multiple loci. 33M72 progenyplants that retain the same degree of phenotypic expression of thesequantitative traits as 33M72 have received significant genotypic andphenotypic contribution from 33M72. This characterization is enhancedwhen such quantitative trait is not exhibited in non-33M72 breedingmaterial used to develop the 33M72 progeny.

As discussed, supra, in addition to phenotypic observations, a plant canalso be described by its genotype. The genotype of a plant can bedescribed through a genetic marker profile which can identify plants ofthe same variety, a related variety or be used to determine or validatea pedigree. Genetic marker profiles can be obtained by techniques suchas Restriction Fragment Length Polymorphisms (RFLPs), Randomly AmplifiedPolymorphic DNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction(AP-PCR), DNA Amplification Fingerprinting (DAF), Sequence CharacterizedAmplified Regions (SCARs), Amplified Fragment Length Polymorphisms(AFLPs), Simple Sequence Repeats (SSRs) which are also referred to asMicrosatellites, and Single Nucleotide Polymorphisms (SNPs). Forexample, see Berry, Don, et al., “Assessing Probability of AncestryUsing Simple Sequence Repeat Profiles: Applications to Maize Hybrids andInbreds”, Genetics, 2002, 161:813–824, which is incorporated byreference herein in its entirety.

Particular markers used for these purposes are not limited to the set ofmarkers disclosed herewithin, but are envisioned to include any type ofmarker and marker profile which provides a means of distinguishingvarieties. In addition to being used for identification of inbredparents, hybrid variety 33M72, a hybrid produced through the use of33M72 or its parents, and the identification or verification of pedigreefor progeny plants produced through the use of 33M72, the genetic markerprofile is also useful in breeding and developing backcross conversions.

Means of performing genetic marker profiles using SSR polymorphisms arewell known in the art. SSRs are genetic markers based on polymorphismsin repeated nucleotide sequences, such as microsatellites. A markersystem based on SSRs can be highly informative in linkage analysisrelative to other marker systems in that multiple alleles may bepresent. Another advantage of this type of marker is that, through useof flanking primers, detection of SSRs can be achieved, for example, bythe polymerase chain reaction (PCR), thereby eliminating the need forlabor-intensive Southern hybridization. The PCR™ detection is done byuse of two oligonucleotide primers flanking the polymorphic segment ofrepetitive DNA. Repeated cycles of heat denaturation of the DNA followedby annealing of the primers to their complementary sequences at lowtemperatures, and extension of the annealed primers with DNA polymerase,comprise the major part of the methodology.

Following amplification, markers can be scored by gel electrophoresis ofthe amplification products. Scoring of marker genotype is based on thesize of the amplified fragment as measured by molecular weight (MW)rounded to the nearest integer. While variation in the primer used or inlaboratory procedures can affect the reported molecular weight, relativevalues should remain constant regardless of the specific primer orlaboratory used. When comparing lines it is preferable if all SSRprofiles are performed in the same lab. An SSR service is available tothe public on a contractual basis by DNA Landmarks inSaint-Jean-sur-Richelieu, Quebec, Canada.

Primers used for the SSRs suggested herein are publicly available andmay be found in the Maize GDB on the World Wide Web at maizegdb.org(sponsored by the USDA Agricultural Research Service), in Sharopova etal. (Plant Mol. Biol. 48(5–6):463–481), Lee et al. (Plant Mol. Biol.48(5–6); 453–461). Primers may be constructed from publicly availablesequence information. Some marker information may be available from DNALandmarks.

A genetic marker profile of a hybrid should be the sum of its inbredparents, e.g., if one inbred parent is homozygous for allele x at aparticular locus, and the other inbred parent is homozygous for allele yat that locus, the F1 hybrid will be x.y (heterozygous) at that locus.The profile can therefore be used to identify the inbred parents ofhybrid 33M72. The determination of the male set of alleles and thefemale set of alleles may be made by profiling the hybrid and thepericarp of the hybrid seed, which is composed of maternal parent cells.The paternal parent profile is obtained by subtracting the pericarpprofile from the hybrid profile.

In addition, plants and plant parts substantially benefiting from theuse of 33M72 in their development such as 33M72 comprising a backcrossconversion, transgene, or genetic sterility factor, may be identified byhaving a molecular marker profile with a high percent identity to 33M72.Such a percent identity might be 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9%identical to 33M72.

The SSR profile of 33M72 also can be used to identify essentiallyderived varieties and other progeny lines developed from the use of33M72, as well as cells and other plant parts thereof. Progeny plantsand plant parts produced using 33M72 may be identified by having amolecular marker profile of at least 25%, 30%, 35%, 40%, 45%, 50%, 55%,60%, 65%, 70%, 75%, 79%, 80%, 81%, 82%, 83%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 99.5% genetic contributionfrom hybrid maize plant 33M72.

Recurrent Selection and Mass Selection

Recurrent selection is a method used in a plant breeding program toimprove a population of plants. 33M72 is suitable for use in a recurrentselection program. The method entails individual plants crosspollinating with each other to form progeny. The progeny are grown andthe superior progeny selected by any number of selection methods, whichinclude individual plant, half-sib progeny, full-sib progeny, selfedprogeny and topcrossing. The selected progeny are cross pollinated witheach other to form progeny for another population. This population isplanted and again superior plants are selected to cross pollinate witheach other. Recurrent selection is a cyclical process and therefore canbe repeated as many times as desired. The objective of recurrentselection is to improve the traits of a population. The improvedpopulation can then be used as a source of breeding material to obtaininbred lines to be used in hybrids or used as parents for a syntheticcultivar. A synthetic cultivar is the resultant progeny formed by theintercrossing of several selected inbreds.

Mass selection is a useful technique when used in conjunction withmolecular marker enhanced selection. In mass selection seeds fromindividuals are selected based on phenotype and/or genotype. Theseselected seeds are then bulked and used to grow the next generation.Bulk selection requires growing a population of plants in a bulk plot,allowing the plants to self-pollinate, harvesting the seed in bulk andthen using a sample of the seed harvested in bulk to plant the nextgeneration. Instead of self pollination, directed pollination could beused as part of the breeding program.

Mutation Breeding

Mutation breeding is one of many methods that could be used to introducenew traits into 33M72. Mutations that occur spontaneously or areartificially induced can be useful sources of variability for a plantbreeder. The goal of artificial mutagenesis is to increase the rate ofmutation for a desired characteristic. Mutation rates can be increasedby many different means including temperature, long-term seed storage,tissue culture conditions, radiation; such as X-rays, Gamma rays (e.g.cobalt 60 or cesium 137), neutrons, (product of nuclear fission byuranium 235 in an atomic reactor), Beta radiation (emitted fromradioisotopes such as phosphorus 32 or carbon 14), or ultravioletradiation (preferably from 2500 to 2900 nm), or chemical mutagens (suchas base analogues (5-bromo-uracil), related compounds (8-ethoxycaffeine), antibiotics (streptonigrin), alkylating agents (sulfurmustards, nitrogen mustards, epoxides, ethylenamines, sulfates,sulfonates, sulfones, lactones), azide, hydroxylamine, nitrous acid, oracridines. Once a desired trait is observed through mutagenesis thetrait may then be incorporated into existing germplasm by traditionalbreeding techniques, such as backcrossing. Details of mutation breedingcan be found in “Principles of Cultivar Development” Fehr, 1993Macmillan Publishing Company the disclosure of which is incorporatedherein by reference. In addition, mutations created in other lines maybe used to produce a backcross conversion of 33M72 that comprises suchmutation.

INDUSTRIAL APPLICABILITY

Maize is used as human food, livestock feed, and as raw material inindustry. The food uses of maize, in addition to human consumption ofmaize kernels, include both products of dry- and wet-milling industries.The principal products of maize dry milling are grits, meal and flour.The maize wet-milling industry can provide maize starch, maize syrups,and dextrose for food use. Maize oil is recovered from maize germ, whichis a by-product of both dry- and wet-milling industries.

Maize, including both grain and non-grain portions of the plant, is alsoused extensively as livestock feed, primarily for beef cattle, dairycattle, hogs, and poultry.

Industrial uses of maize include production of ethanol, maize starch inthe wet-milling industry and maize flour in the dry-milling industry.The industrial applications of maize starch and flour are based onfunctional properties, such as viscosity, film formation, adhesiveproperties, and ability to suspend particles. The maize starch and flourhave application in the paper and textile industries. Other industrialuses include applications in adhesives, building materials, foundrybinders, laundry starches, explosives, oil-well muds, and other miningapplications.

Plant parts other than the grain of maize are also used in industry: forexample, stalks and husks are made into paper and wallboard and cobs areused for fuel and to make charcoal.

The seed of the hybrid maize plant, the plant produced from the seed, aplant produced from crossing of maize hybrid plant 33M72 and variousparts of the hybrid maize plant and transgenic versions of theforegoing, can be utilized for human food, livestock feed, and as a rawmaterial in industry.

DEPOSITS

Applicants have made a deposit of at least 2500 seeds of hybrid maize33M72 with the American Type Culture Collection (ATCC), Manassas, Va.20110 USA, ATCC Deposit No. PTA-5759. The seeds deposited with the ATCCon Jan. 12, 2004 were taken from the deposit maintained by PioneerHi-Bred International, Inc., 7100 NW 62nd Avenue, Johnston, Iowa50131-1000 since prior to the filing date of this application. Access tothis deposit will be available during the pendency of the application tothe Commissioner of Patents and Trademarks and persons determined by theCommissioner to be entitled thereto upon request. Upon allowance of anyclaims in the application, the Applicants will make available to thepublic, pursuant to 37 C.F.R. § 1.808, sample(s) of the deposit of atleast 2500 seeds of hybrid maize 33M72 with the American Type CultureCollection (ATCC), 10801 University Boulevard, Manassas, Va. 20110-2209.This deposit of seed of hybrid maize 33M72 will be maintained in theATCC depository, which is a public depository, for a period of 30 years,or 5 years after the most recent request, or for the enforceable life ofthe patent, whichever is longer, and will be replaced if it becomesnonviable during that period. Additionally, Applicants have satisfiedall the requirements of 37 C.F.R. §§1.801–1.809, including providing anindication of the viability of the sample upon deposit. Applicants haveno authority to waive any restrictions imposed by law on the transfer ofbiological material or its transportation in commerce. Applicants do notwaive any infringement of their rights granted under this patent orrights applicable to Hybrid Maize 33M72 under the Plant VarietyProtection Act (7 USC 2321 et seq.).

All publications, patents and patent applications mentioned in thespecification are indicative of the level of those skilled in the art towhich this invention pertains. All such publications, patents and patentapplications are incorporated by reference herein to the same extent asif each was specifically and individually indicated to be incorporatedby reference herein.

The foregoing invention has been described in detail by way ofillustration and example for purposes of clarity and understanding.However, it will be obvious that certain changes and modifications suchas backcross conversions and mutations, somoclonal variants, variantindividuals selected from large populations of the plants of the instantinbred and the like may be practiced within the scope of the invention,as limited only by the scope of the appended claims.

1. A seed of hybrid maize variety designated 33M72, representative seedof said variety having been deposited under ATCC Accession NumberPTA-5759.
 2. A maize plant, or a part thereof, produced by growing theseed of claim
 1. 3. Pollen of the plant of claim
 2. 4. An ovule orovules of the plant of claim
 2. 5. A maize plant, or a part thereof,having all the physiological and morphological characteristics of thehybrid maize variety 33M72, representative seed of said variety havingbeen deposited under ATCC Accession No: PTA-5759.
 6. A tissue culture ofregenerable cells produced from the plant of claim
 2. 7. Protoplasts orcallus produced from the tissue culture of claim
 6. 8. The tissueculture of claim 6, wherein the regenerable cells of the tissue cultureare produced from protoplasts or from tissue of a plant part selectedfrom the group consisting of pollen, embryo, immature embryo,meristematic cells, immature tassels, microspores, root, root tip,anther, silk, flower, kernel, ear, cob, husk and stalk.
 9. A maize plantregenerated from the tissue culture of claim 6, said plant having allthe morphological and physiological characteristics of hybrid maizevariety 33M72, representative seed of said variety having been depositedunder ATCC Accession No. PTA-5759.
 10. A process for producing an F1hybrid maize seed, said method comprising crossing the plant of claim 2with a different maize plant and harvesting the F1 hybrid maize seed.11. The process of claim 10, further comprising growing the F1 hybridmaize seed to produce a hybrid maize plant.
 12. A process for producinga maize seed, comprising crossing the plant of claim 2 with itself or adifferent maize plant and harvesting the resultant maize seed.